Uses of kinase inhibitors for inducing and maintaining pluripotency

ABSTRACT

The present disclosure provides compounds of any one of Formulae (A) to (L). The present disclosure also provides compositions, uses, and methods that include or involve a compound described herein, a serine/threonine-protein kinase B-Raf (BRAF) inhibitor, an epidermal growth factor receptor (EGFR) inhibitor, a vascular endothelial growth factor 1 (VEGFR1) inhibitor, a fibroblast growth factor receptor 1 (FGFR1) inhibitor, or a combination thereof. The compounds, compositions, uses, and methods are useful in changing the pluripotency state of a vertebrate cell to a more naive state.

RELATED APPLICATIONS

This application claims priority under 35 U.S.C. §119(e) to U.S.provisional applications, U.S. Ser. No. 62/014,674, filed Jun. 19, 2014,and U.S. Ser. No. 62/045,337, filed Sep. 3, 2014, each of which isincorporated herein by reference.

GOVERNMENT SUPPORT

This invention was made with U.S. Government support under grant numberR01-CA084198 awarded by the National Institutes of Health. The U.S.Government has certain rights in the invention.

BACKGROUND OF THE INVENTION

Human pluripotent stem cells, including embryonic stem cells (ESCs) andinduced pluripotent stem cells (iPSCs), hold great promise forregenerative medicine and disease modeling (Hanna et al., 2010b). Fullrealization of their potential is currently constrained by laboriousculture requirements and inconsistencies in developmental potentialbetween lines (Melichar et al., 2011; Osafune et al., 2008). Researchershave had a relatively easy time genetically manipulating and preventingdifferentiation in mouse ES and iPS cells. However, human ES cells canbe more technically demanding to culture and exhibit properties such asslow growth and poor tolerance to passaging as single cells. Thus, thereis a need for more effective techniques to isolate and culture humanpluripotent stem cells.

SUMMARY OF THE INVENTION

One aspect of the disclosure provides methods for changing thepluripotency state of a vertebrate cell to a more nave state, themethods comprising: culturing a pluripotent vertebrate cell in thepresence of a serine/threonine-protein kinase B-Raf (BRAF) inhibitor, anepidermal growth factor receptor (EGFR) inhibitor, a vascularendothelial growth factor 1 (VEGFR1) inhibitor, or a fibroblast growthfactor receptor 1 (FGFR1) inhibitor; and maintaining the cell in cultureunder conditions suitable and a time sufficient to convert thepluripotency state of the vertebrate cell to a more nave state than thepluripotency state of the vertebrate cell of culturing step.

Another aspect of the disclosure provides methods for changing thepluripotency state of a vertebrate cell to a more nave state, the methodcomprising: culturing a pluripotent vertebrate cell in the presence of acompound of any one of Formulae (A) to (L):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof; and maintaining the cell in culture underconditions suitable and a time sufficient to convert the pluripotencystate of the vertebrate cell to a more naïve state than the pluripotencystate of the vertebrate cell of culturing step. In some embodiments, thetime sufficient to convert the pluripotency state of the vertebrate cellto a more naïve state is at least about 5 days (e.g., about 10 days).

Exemplary compounds useful in a system, composition, kit, or methoddescribed herein include, but are not limited to, AMG706, AMN-107,AZ-628, BAY 73-4506, BAY-439006, GDC-0879, KIN001-260, SB590885,CHIR99021, KIN001-244, KU55933, SP600125, PD0332991, PD173074, WZ-7043,BIBF-1120, PHA-665752, SU11248, SU11274, KIN001-220, WH-4-023, WH-4-025,XMD 8-92, XMD11-50, XMD 8-85, IM12, PD0325901, and Y-27632.

Another aspect of the disclosure provides compositions comprising naïvepluripotent vertebrate cells produced by a method described herein.

Another aspect of the disclosure provides naïve pluripotent vertebratecells, wherein the cells have a global gene expression profile whichclusters with naïve mouse ESCs as opposed to stem cell lines derivedfrom mouse epiblast (EpiSCs) and/or less naïve human ESCs.

Yet another aspect of the disclosure relates to naïve pluripotentvertebrate cells, wherein the cells have a global gene expressionprofile which clusters with naïve mouse ESCs as opposed to stem celllines derived from mouse epiblast (EpiSCs) and/or less naïve human ESCs.

Another aspect of the disclosure provides kits for changing thepluripotency state of a vertebrate cell to a more naïve state, the kitscomprising a pluripotent vertebrate cell; and cell culture mediumcomprising a serine/threonine-protein kinase B-Raf (BRAF) inhibitor, anepidermal growth factor receptor (EGFR) inhibitor, a vascularendothelial growth factor 1 (VEGFR1) inhibitor, or a fibroblast growthfactor receptor 1 (FGFR1) inhibitor.

In yet another aspect, the present disclosure provides compounds,compositions, and kits described herein for use in a method of thepresent disclosure.

The details of particular embodiments of the invention are set forthherein. Other features, objects, and advantages of the invention will beapparent from the Detailed Description, the Figures, the Examples, andthe Claims.

DEFINITIONS

Definitions of specific functional groups and chemical terms aredescribed in more detail below. The chemical elements are identified inaccordance with the Periodic Table of the Elements, CAS version,Handbook of Chemistry and Physics, 75^(th) Ed., inside cover, andspecific functional groups are generally defined as described therein.Additionally, general principles of organic chemistry, as well asspecific functional moieties and reactivity, are described in OrganicChemistry, Thomas Sorrell, University Science Books, Sausalito, 1999;Smith and March March's Advanced Organic Chemistry, 5^(th) Edition, JohnWiley & Sons, Inc., New York, 2001; Larock, Comprehensive OrganicTransformations, VCH Publishers, Inc., New York, 1989; and Carruthers,Some Modern Methods of Organic Synthesis, 3^(rd) Edition, CambridgeUniversity Press, Cambridge, 1987.

Compounds described herein can comprise one or more asymmetric centers,and thus can exist in various stereoisomeric forms, e.g., enantiomersand/or diastereomers. For example, the compounds described herein can bein the form of an individual enantiomer, diastereomer or geometricisomer, or can be in the form of a mixture of stereoisomers, includingracemic mixtures and mixtures enriched in one or more stereoisomer.Isomers can be isolated from mixtures by methods known to those skilledin the art, including chiral high pressure liquid chromatography (HPLC)and the formation and crystallization of chiral salts; or preferredisomers can be prepared by asymmetric syntheses. See, for example,Jacques et al., Enantiomers, Racemates and Resolutions (WileyInterscience, New York, 1981); Wilen et al., Tetrahedron 33:2725 (1977);Eliel, E. L. Stereochemistry of Carbon Compounds (McGrawHill, NY, 1962);and Wilen, S. H. Tables of Resolving Agents and Optical Resolutions p.268 (E. L. Eliel, Ed., Univ. of Notre Dame Press, Notre Dame, Ind.1972). The disclosure additionally encompasses compounds as individualisomers substantially free of other isomers, and alternatively, asmixtures of various isomers.

In a formula, --- is absent or a single bond, and

, or

is a single or double bond.

The term “heteroatom” refers to an atom that is not hydrogen or carbon.In certain embodiments, the heteroatom is nitrogen. In certainembodiments, the heteroatom is oxygen. In certain embodiments, theheteroatom is sulfur.

When a range of values is listed, it is intended to encompass each valueand sub range within the range. For example “C₁₋₆ alkyl” is intended toencompass, C₁, C₂, C₃, C₄, C₅, C₆, C₁₋₆, C₁₋₅, C₁₋₄, C₁₋₃, C₁₋₂, C₂₋₆,C₂₋₅, C₂₋₄, C₂₋₃, C₃₋₆, C₃₋₅, C₃₋₄, C₄₋₆, C₄₋₅, and C₅₋₆ alkyl.

The term “aliphatic” refers to alkyl, alkenyl, alkynyl, and carbocyclicgroups. Likewise, the term “heteroaliphatic” refers to heteroalkyl,heteroalkenyl, heteroalkynyl, and heterocyclic groups.

The term “alkyl” refers to a radical of a straightchain or branchedsaturated hydrocarbon group having from 1 to 10 carbon atoms (“C₁₋₁₀alkyl”). In some embodiments, an alkyl group has 1 to 9 carbon atoms(“C₁₋₉ alkyl”). In some embodiments, an alkyl group has 1 to 8 carbonatoms (“C₁₋₈ alkyl”). In some embodiments, an alkyl group has 1 to 7carbon atoms (“C₁₋₇ alkyl”). In some embodiments, an alkyl group has 1to 6 carbon atoms (“C₁₋₆ alkyl”). In some embodiments, an alkyl grouphas 1 to 5 carbon atoms (“C₁₋₅ alkyl”). In some embodiments, an alkylgroup has 1 to 4 carbon atoms (“C₁₋₄ alkyl”). In some embodiments, analkyl group has 1 to 3 carbon atoms (“C₁₋₃ alkyl”). In some embodiments,an alkyl group has 1 to 2 carbon atoms (“C₁₋₂ alkyl”). In someembodiments, an alkyl group has 1 carbon atom (“C₁ alkyl”). In someembodiments, an alkyl group has 2 to 6 carbon atoms (“C₂₋₆ alkyl”).Examples of C₁₋₆ alkyl groups include methyl (C₁), ethyl (C₂), n-propyl(C₃), isopropyl (C₃), n-butyl (C₄), tert-butyl (C₄), sec-butyl (C₄),iso-butyl (C₄), n-pentyl (C₅), 3-pentanyl (C₅), amyl (C₅), neopentyl(C₅), 3-methyl-2-butanyl (C₅), tertiary amyl (C₅), and n-hexyl (C₆).Additional examples of alkyl groups include n-heptyl (C₇), n-octyl (C₈)and the like. Unless otherwise specified, each instance of an alkylgroup is independently unsubstituted (an “unsubstituted alkyl”) orsubstituted (a “substituted alkyl”) with one or more substituents. Incertain embodiments, the alkyl group is an unsubstituted C₁₋₁₀ alkyl(e.g., —CH₃). In certain embodiments, the alkyl group is a substitutedC₁₋₁₀ alkyl.

The term “haloalkyl” is a substituted alkyl group, wherein one or moreof the hydrogen atoms are independently replaced by a halogen, e.g.,fluoro, bromo, chloro, or iodo. “Perhaloalkyl” is a subset of haloalkyl,and refers to an alkyl group wherein all of the hydrogen atoms areindependently replaced by a halogen, e.g., fluoro, bromo, chloro, oriodo. In some embodiments, the haloalkyl moiety has 1 to 8 carbon atoms(“C₁₋₈ haloalkyl”). In some embodiments, the haloalkyl moiety has 1 to 6carbon atoms (“C₁₋₆ haloalkyl”). In some embodiments, the haloalkylmoiety has 1 to 4 carbon atoms (“C₁₋₄ haloalkyl”). In some embodiments,the haloalkyl moiety has 1 to 3 carbon atoms (“C₁₋₃ haloalkyl”). In someembodiments, the haloalkyl moiety has 1 to 2 carbon atoms (“C₁₋₂haloalkyl”). In some embodiments, all of the haloalkyl hydrogen atomsare replaced with fluoro to provide a perfluoroalkyl group. In someembodiments, all of the haloalkyl hydrogen atoms are replaced withchloro to provide a “perchloroalkyl” group. Examples of haloalkyl groupsinclude —CF₃, —CF₂CF₃, —CF₂CF₂CF₃, —CCl₃, —CFCl₂, —CF₂Cl, and the like.

The term “heteroalkyl” refers to an alkyl group, which further includesat least one heteroatom (e.g., 1, 2, 3, or 4 heteroatoms) selected fromoxygen, nitrogen, or sulfur within (i.e., inserted between adjacentcarbon atoms of) and/or placed at one or more terminal position(s) ofthe parent chain. In certain embodiments, a heteroalkyl group refers toa saturated group having from 1 to 10 carbon atoms and 1 or moreheteroatoms within the parent chain (“heteroC₁₋₁₀ alkyl”). In someembodiments, a heteroalkyl group is a saturated group having 1 to 9carbon atoms and 1 or more heteroatoms within the parent chain(“heteroC₁₋₉ alkyl”). In some embodiments, a heteroalkyl group is asaturated group having 1 to 8 carbon atoms and 1 or more heteroatomswithin the parent chain (“heteroC₁₋₈ alkyl”). In some embodiments, aheteroalkyl group is a saturated group having 1 to 7 carbon atoms and 1or more heteroatoms within the parent chain (“heteroC₁₋₇ alkyl”). Insome embodiments, a heteroalkyl group is a saturated group having 1 to 6carbon atoms and 1 or more heteroatoms within the parent chain(“heteroC₁₋₆ alkyl”). In some embodiments, a heteroalkyl group is asaturated group having 1 to 5 carbon atoms and 1 or 2 heteroatoms withinthe parent chain (“heteroC₁₋₅ alkyl”). In some embodiments, aheteroalkyl group is a saturated group having 1 to 4 carbon atoms andfor 2 heteroatoms within the parent chain (“heteroC₁₋₄ alkyl”). In someembodiments, a heteroalkyl group is a saturated group having 1 to 3carbon atoms and 1 heteroatom within the parent chain (“heteroC₁₋₃alkyl”). In some embodiments, a heteroalkyl group is a saturated grouphaving 1 to 2 carbon atoms and 1 heteroatom within the parent chain(“heteroC₁₋₂ alkyl”). In some embodiments, a heteroalkyl group is asaturated group having 1 carbon atom and 1 heteroatom (“heteroC₁alkyl”). In some embodiments, a heteroalkyl group is a saturated grouphaving 2 to 6 carbon atoms and 1 or 2 heteroatoms within the parentchain (“heteroC₂₋₆ alkyl”). Unless otherwise specified, each instance ofa heteroalkyl group is independently unsubstituted (an “unsubstitutedheteroalkyl”) or substituted (a “substituted heteroalkyl”) with one ormore substituents. In certain embodiments, the heteroalkyl group is anunsubstituted heteroC₁₋₁₀ alkyl. In certain embodiments, the heteroalkylgroup is a substituted heteroC₁₋₁₀ alkyl.

The term “alkenyl” refers to a radical of a straight-chain or branchedhydrocarbon group having from 2 to 10 carbon atoms and one or morecarbon-carbon double bonds (e.g., 1, 2, 3, or 4 double bonds). In someembodiments, an alkenyl group has 2 to 9 carbon atoms (“C₂₋₉ alkenyl”).In some embodiments, an alkenyl group has 2 to 8 carbon atoms (“C₂₋₈alkenyl”). In some embodiments, an alkenyl group has 2 to 7 carbon atoms(“C₂₋₇ alkenyl”). In some embodiments, an alkenyl group has 2 to 6carbon atoms (“C₂₋₆ alkenyl”). In some embodiments, an alkenyl group has2 to 5 carbon atoms (“C₂₋₅ alkenyl”). In some embodiments, an alkenylgroup has 2 to 4 carbon atoms (“C₂₋₄ alkenyl”). In some embodiments, analkenyl group has 2 to 3 carbon atoms (“C₂₋₃ alkenyl”). In someembodiments, an alkenyl group has 2 carbon atoms (“C₂ alkenyl”). The oneor more carbon-carbon double bonds can be internal (such as in2-butenyl) or terminal (such as in 1-butenyl). Examples of C₂₋₄ alkenylgroups include ethenyl (C₂), 1-propenyl (C₃), 2-propenyl (C₃), 1-butenyl(C₄), 2-butenyl (C₄), butadienyl (C₄), and the like. Examples of C₂₋₆alkenyl groups include the aforementioned C₂₋₄ alkenyl groups as well aspentenyl (C₅), pentadienyl (C₅), hexenyl (C₆), and the like. Additionalexamples of alkenyl include heptenyl (C₇), octenyl (C₈), octatrienyl(C₈), and the like. Unless otherwise specified, each instance of analkenyl group is independently unsubstituted (an “unsubstitutedalkenyl”) or substituted (a “substituted alkenyl”) with one or moresubstituents. In certain embodiments, the alkenyl group is anunsubstituted C₂₋₁₀ alkenyl. In certain embodiments, the alkenyl groupis a substituted C₂₋₁₀ alkenyl. In an alkenyl group, a C═C double bondfor which the stereochemistry is unspecified (e.g., —CH═CHCH₃ or

may be an (E)- or (Z)-double bond.

The term “heteroalkenyl” refers to an alkenyl group, which furtherincludes at least one heteroatom (e.g., 1, 2, 3, or 4 heteroatoms)selected from oxygen, nitrogen, or sulfur within (i.e., inserted betweenadjacent carbon atoms of) and/or placed at one or more terminalposition(s) of the parent chain. In certain embodiments, a heteroalkenylgroup refers to a group having from 2 to 10 carbon atoms, at least onedouble bond, and 1 or more heteroatoms within the parent chain(“heteroC₂₋₁₀ alkenyl”). In some embodiments, a heteroalkenyl group has2 to 9 carbon atoms at least one double bond, and 1 or more heteroatomswithin the parent chain (“heteroC₂₋₉ alkenyl”). In some embodiments, aheteroalkenyl group has 2 to 8 carbon atoms, at least one double bond,and 1 or more heteroatoms within the parent chain (“heteroC₂₋₈alkenyl”). In some embodiments, a heteroalkenyl group has 2 to 7 carbonatoms, at least one double bond, and 1 or more heteroatoms within theparent chain (“heteroC₂₋₇ alkenyl”). In some embodiments, aheteroalkenyl group has 2 to 6 carbon atoms, at least one double bond,and 1 or more heteroatoms within the parent chain (“heteroC₂₋₆alkenyl”). In some embodiments, a heteroalkenyl group has 2 to 5 carbonatoms, at least one double bond, and 1 or 2 heteroatoms within theparent chain (“heteroC₂₋₅ alkenyl”). In some embodiments, aheteroalkenyl group has 2 to 4 carbon atoms, at least one double bond,and for 2 heteroatoms within the parent chain (“heteroC₂₋₄ alkenyl”). Insome embodiments, a heteroalkenyl group has 2 to 3 carbon atoms, atleast one double bond, and 1 heteroatom within the parent chain(“heteroC₂₋₃ alkenyl”). In some embodiments, a heteroalkenyl group has 2to 6 carbon atoms, at least one double bond, and 1 or 2 heteroatomswithin the parent chain (“heteroC₂₋₆ alkenyl”). Unless otherwisespecified, each instance of a heteroalkenyl group is independentlyunsubstituted (an “unsubstituted heteroalkenyl”) or substituted (a“substituted heteroalkenyl”) with one or more substituents. In certainembodiments, the heteroalkenyl group is an unsubstituted heteroC₂₋₁₀alkenyl. In certain embodiments, the heteroalkenyl group is asubstituted heteroC₂₋₁₀ alkenyl.

The term “alkynyl” refers to a radical of a straight-chain or branchedhydrocarbon group having from 2 to 10 carbon atoms and one or morecarbon-carbon triple bonds (e.g., 1, 2, 3, or 4 triple bonds) (“C₂₋₁₀alkynyl”). In some embodiments, an alkynyl group has 2 to 9 carbon atoms(“C₂₋₉ alkynyl”). In some embodiments, an alkynyl group has 2 to 8carbon atoms (“C₂₋₈ alkynyl”). In some embodiments, an alkynyl group has2 to 7 carbon atoms (“C₂₋₇ alkynyl”). In some embodiments, an alkynylgroup has 2 to 6 carbon atoms (“C₂₋₆ alkynyl”). In some embodiments, analkynyl group has 2 to 5 carbon atoms (“C₂₋₅ alkynyl”). In someembodiments, an alkynyl group has 2 to 4 carbon atoms (“C₂₋₄ alkynyl”).In some embodiments, an alkynyl group has 2 to 3 carbon atoms (“C₂₋₃alkynyl”). In some embodiments, an alkynyl group has 2 carbon atoms (“C₂alkynyl”). The one or more carbon carbon triple bonds can be internal(such as in 2-butynyl) or terminal (such as in 1-butynyl). Examples ofC₂₋₄ alkynyl groups include, without limitation, ethynyl (C₂),1-propynyl (C₃), 2-propynyl (C₃), 1-butynyl (C₄), 2-butynyl (C₄), andthe like. Examples of C₂₋₆ alkenyl groups include the aforementionedC₂₋₄ alkynyl groups as well as pentynyl (C₅), hexynyl (C₆), and thelike. Additional examples of alkynyl include heptynyl (C₇), octynyl(C₈), and the like. Unless otherwise specified, each instance of analkynyl group is independently unsubstituted (an “unsubstitutedalkynyl”) or substituted (a “substituted alkynyl”) with one or moresubstituents. In certain embodiments, the alkynyl group is anunsubstituted C₂₋₁₀ alkynyl. In certain embodiments, the alkynyl groupis a substituted C₂₋₁₀ alkynyl.

The term “heteroalkynyl” refers to an alkynyl group, which furtherincludes at least one heteroatom (e.g., 1, 2, 3, or 4 heteroatoms)selected from oxygen, nitrogen, or sulfur within (i.e., inserted betweenadjacent carbon atoms of) and/or placed at one or more terminalposition(s) of the parent chain. In certain embodiments, a heteroalkynylgroup refers to a group having from 2 to 10 carbon atoms, at least onetriple bond, and 1 or more heteroatoms within the parent chain(“heteroC₂₋₁₀ alkynyl”). In some embodiments, a heteroalkynyl group has2 to 9 carbon atoms, at least one triple bond, and 1 or more heteroatomswithin the parent chain (“heteroC₂₋₉ alkynyl”). In some embodiments, aheteroalkynyl group has 2 to 8 carbon atoms, at least one triple bond,and 1 or more heteroatoms within the parent chain (“heteroC₂₋₈alkynyl”). In some embodiments, a heteroalkynyl group has 2 to 7 carbonatoms, at least one triple bond, and 1 or more heteroatoms within theparent chain (“heteroC₂₋₇ alkynyl”). In some embodiments, aheteroalkynyl group has 2 to 6 carbon atoms, at least one triple bond,and 1 or more heteroatoms within the parent chain (“heteroC₂₋₆alkynyl”). In some embodiments, a heteroalkynyl group has 2 to 5 carbonatoms, at least one triple bond, and 1 or 2 heteroatoms within theparent chain (“heteroC₂₋₅ alkynyl”). In some embodiments, aheteroalkynyl group has 2 to 4 carbon atoms, at least one triple bond,and for 2 heteroatoms within the parent chain (“heteroC₂₋₄ alkynyl”). Insome embodiments, a heteroalkynyl group has 2 to 3 carbon atoms, atleast one triple bond, and 1 heteroatom within the parent chain(“heteroC₂₋₃ alkynyl”). In some embodiments, a heteroalkynyl group has 2to 6 carbon atoms, at least one triple bond, and 1 or 2 heteroatomswithin the parent chain (“heteroC₂₋₆ alkynyl”). Unless otherwisespecified, each instance of a heteroalkynyl group is independentlyunsubstituted (an “unsubstituted heteroalkynyl”) or substituted (a“substituted heteroalkynyl”) with one or more substituents. In certainembodiments, the heteroalkynyl group is an unsubstituted heteroC₂₋₁₀alkynyl. In certain embodiments, the heteroalkynyl group is asubstituted heteroC₂₋₁₀ alkynyl.

The term “carbocyclyl” or “carbocyclic” refers to a radical of anonaromatic cyclic hydrocarbon group having from 3 to 14 ring carbonatoms (“C₃₋₁₄ carbocyclyl”) and zero heteroatoms in the nonaromatic ringsystem. In some embodiments, a carbocyclyl group has 3 to 10 ring carbonatoms (“C₃₋₁₀ carbocyclyl”). In some embodiments, a carbocyclyl grouphas 3 to 8 ring carbon atoms (“C₃₋₈ carbocyclyl”). In some embodiments,a carbocyclyl group has 3 to 7 ring carbon atoms (“C₃₋₇ carbocyclyl”).In some embodiments, a carbocyclyl group has 3 to 6 ring carbon atoms(“C₃₋₆ carbocyclyl”). In some embodiments, a carbocyclyl group has 4 to6 ring carbon atoms (“C₄₋₆ carbocyclyl”). In some embodiments, acarbocyclyl group has 5 to 6 ring carbon atoms (“C₅₋₆ carbocyclyl”). Insome embodiments, a carbocyclyl group has 5 to 10 ring carbon atoms(“C₅₋₁₀ carbocyclyl”). Exemplary C₃₋₆ carbocyclyl groups include,without limitation, cyclopropyl (C₃), cyclopropenyl (C₃), cyclobutyl(C₄), cyclobutenyl (C₄), cyclopentyl (C₅), cyclopentenyl (C₅),cyclohexyl (C₆), cyclohexenyl (C₆), cyclohexadienyl (C₆), and the like.Exemplary C₃₋₈ carbocyclyl groups include, without limitation, theaforementioned C₃₋₆ carbocyclyl groups as well as cycloheptyl (C₇),cycloheptenyl (C₇), cycloheptadienyl (C₇), cycloheptatrienyl (C₇),cyclooctyl (C₈), cyclooctenyl (C₈), bicyclo[2.2.1]heptanyl (C₇),bicyclo[2.2.2]octanyl (C₈), and the like. Exemplary C₃₋₁₀ carbocyclylgroups include, without limitation, the aforementioned C₃₋₈ carbocyclylgroups as well as cyclononyl (C₉), cyclononenyl (C₉), cyclodecyl (C₁₀),cyclodecenyl (C₁₀), octahydro-1H-indenyl (C₉), decahydronaphthalenyl(C₁₀), spiro[4.5]decanyl (C₁₀), and the like. As the foregoing examplesillustrate, in certain embodiments, the carbocyclyl group is eithermonocyclic (“monocyclic carbocyclyl”) or polycyclic (e.g., containing afused, bridged or spiro ring system such as a bicyclic system (“bicycliccarbocyclyl”) or tricyclic system (“tricyclic carbocyclyl”)) and can besaturated or can contain one or more carbon-carbon double or triplebonds. “Carbocyclyl” also includes ring systems wherein the carbocyclylring, as defined above, is fused with one or more aryl or heteroarylgroups wherein the point of attachment is on the carbocyclyl ring, andin such instances, the number of carbons continue to designate thenumber of carbons in the carbocyclic ring system. Unless otherwisespecified, each instance of a carbocyclyl group is independentlyunsubstituted (an “unsubstituted carbocyclyl”) or substituted (a“substituted carbocyclyl”) with one or more substituents. In certainembodiments, the carbocyclyl group is an unsubstituted C₃₋₁₄carbocyclyl. In certain embodiments, the carbocyclyl group is asubstituted C₃₋₁₄ carbocyclyl. In some embodiments, carbocyclyl is amonocyclic, saturated carbocyclyl group having from 3 to 14 ring carbonatoms (“C₃₋₁₄ cycloalkyl”). In some embodiments, a cycloalkyl group has3 to 10 ring carbon atoms (“C₃₋₁₀ cycloalkyl”). In some embodiments, acycloalkyl group has 3 to 8 ring carbon atoms (“C₃₋₈ cycloalkyl”). Insome embodiments, a cycloalkyl group has 3 to 6 ring carbon atoms (“C₃₋₆cycloalkyl”). In some embodiments, a cycloalkyl group has 4 to 6 ringcarbon atoms (“C₄₋₆ cycloalkyl”). In some embodiments, a cycloalkylgroup has 5 to 6 ring carbon atoms (“C₅₋₆ cycloalkyl”). In someembodiments, a cycloalkyl group has 5 to 10 ring carbon atoms (“C₅₋₁₀cycloalkyl”). Examples of C₅₋₆ cycloalkyl groups include cyclopentyl(C₅) and cyclohexyl (C₅). Examples of C₃₋₆ cycloalkyl groups include theaforementioned C₅₋₆ cycloalkyl groups as well as cyclopropyl (C₃) andcyclobutyl (C₄). Examples of C₃₋₈ cycloalkyl groups include theaforementioned C₃₋₆ cycloalkyl groups as well as cycloheptyl (C₇) andcyclooctyl (C₈). Unless otherwise specified, each instance of acycloalkyl group is independently unsubstituted (an “unsubstitutedcycloalkyl”) or substituted (a “substituted cycloalkyl”) with one ormore substituents. In certain embodiments, the cycloalkyl group is anunsubstituted C₃₋₁₄ cycloalkyl. In certain embodiments, the cycloalkylgroup is a substituted C₃₋₁₄ cycloalkyl.

The term “heterocyclyl” or “heterocyclic” refers to a radical of a 3 to14-membered nonaromatic ring system having ring carbon atoms and 1 to 4ring heteroatoms, wherein each heteroatom is independently selected fromnitrogen, oxygen, and sulfur (“3-14 membered heterocyclyl”). Inheterocyclyl groups that contain one or more nitrogen atoms, the pointof attachment can be a carbon or nitrogen atom, as valency permits. Aheterocyclyl group can either be monocyclic (“monocyclic heterocyclyl”)or polycyclic (e.g., a fused, bridged or spiro ring system such as abicyclic system (“bicyclic heterocyclyl”) or tricyclic system(“tricyclic heterocyclyl”)), and can be saturated or can contain one ormore carbon carbon double or triple bonds. Heterocyclyl polycyclic ringsystems can include one or more heteroatoms in one or both rings.“Heterocyclyl” also includes ring systems wherein the heterocyclyl ring,as defined above, is fused with one or more carbocyclyl groups whereinthe point of attachment is either on the carbocyclyl or heterocyclylring, or ring systems wherein the heterocyclyl ring, as defined above,is fused with one or more aryl or heteroaryl groups, wherein the pointof attachment is on the heterocyclyl ring, and in such instances, thenumber of ring members continue to designate the number of ring membersin the heterocyclyl ring system. Unless otherwise specified, eachinstance of heterocyclyl is independently unsubstituted (an“unsubstituted heterocyclyl”) or substituted (a “substitutedheterocyclyl”) with one or more substituents. In certain embodiments,the heterocyclyl group is an unsubstituted 3-14-membered heterocyclyl.In certain embodiments, the heterocyclyl group is a substituted3-14-membered heterocyclyl.

In some embodiments, a heterocyclyl group is a 5-10-membered nonaromaticring system having ring carbon atoms and 1-4 ring heteroatoms, whereineach heteroatom is independently selected from nitrogen, oxygen, andsulfur (“5-10-membered heterocyclyl”). In some embodiments, aheterocyclyl group is a 5-8-membered nonaromatic ring system having ringcarbon atoms and 1-4 ring heteroatoms, wherein each heteroatom isindependently selected from nitrogen, oxygen, and sulfur (“5-8-memberedheterocyclyl”). In some embodiments, a heterocyclyl group is a5-6-membered nonaromatic ring system having ring carbon atoms and 1-4ring heteroatoms, wherein each heteroatom is independently selected fromnitrogen, oxygen, and sulfur (“5-6-membered heterocyclyl”). In someembodiments, the 5-6-membered heterocyclyl has 1-3 ring heteroatomsselected from nitrogen, oxygen, and sulfur. In some embodiments, the5-6-membered heterocyclyl has 1-2 ring heteroatoms selected fromnitrogen, oxygen, and sulfur. In some embodiments, the 5-6 memberedheterocyclyl has 1 ring heteroatom selected from nitrogen, oxygen, andsulfur.

Exemplary 3-membered heterocyclyl groups containing 1 heteroatominclude, without limitation, azirdinyl, oxiranyl, and thiiranyl.Exemplary 4-membered heterocyclyl groups containing 1 heteroatominclude, without limitation, azetidinyl, oxetanyl, and thietanyl.Exemplary 5-membered heterocyclyl groups containing 1 heteroatominclude, without limitation, tetrahydrofuranyl, dihydrofuranyl,tetrahydrothiophenyl, dihydrothiophenyl, pyrrolidinyl, dihydropyrrolyl,and pyrrolyl-2,5-dione. Exemplary 5 membered heterocyclyl groupscontaining 2 heteroatoms include, without limitation, dioxolanyl,oxathiolanyl and dithiolanyl. Exemplary 5-membered heterocyclyl groupscontaining 3 heteroatoms include, without limitation, triazolinyl,oxadiazolinyl, and thiadiazolinyl. Exemplary 6-membered heterocyclylgroups containing 1 heteroatom include, without limitation, piperidinyl,tetrahydropyranyl, dihydropyridinyl, and thianyl. Exemplary 6-memberedheterocyclyl groups containing 2 heteroatoms include, withoutlimitation, piperazinyl, morpholinyl, dithianyl, and dioxanyl. Exemplary6-membered heterocyclyl groups containing 2 heteroatoms include, withoutlimitation, triazinanyl. Exemplary 7-membered heterocyclyl groupscontaining 1 heteroatom include, without limitation, azepanyl, oxepanyland thiepanyl. Exemplary 8-membered heterocyclyl groups containing 1heteroatom include, without limitation, azocanyl, oxecanyl andthiocanyl. Exemplary bicyclic heterocyclyl groups include, withoutlimitation, indolinyl, isoindolinyl, dihydrobenzofuranyl,dihydrobenzothienyl, tetrahydrobenzothienyl, tetrahydrobenzofuranyl,tetrahydroindolyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl,decahydroquinolinyl, decahydroisoquinolinyl, octahydrochromenyl,octahydroisochromenyl, decahydronaphthyridinyl,decahydro-1,8-naphthyridinyl, octahydropyrrolo[3,2-b]pyrrole, indolinyl,phthalimidyl, naphthalimidyl, chromanyl, chromenyl,1H-benzo[e][1,4]diazepinyl, 1,4,5,7-tetrahydropyrano[3,4-b]pyrrolyl,5,6-dihydro-4H-furo[3,2-b]pyrrolyl, 6,7-dihydro-5H-furo[3,2-b]pyranyl,5,7-dihydro-4H-thieno[2,3-c]pyranyl,2,3-dihydro-1H-pyrrolo[2,3-b]pyridinyl, 2,3-dihydrofuro[2,3-b]pyridinyl,4,5,6,7-tetrahydro-1H-pyrrolo[2,3-b]pyridinyl,4,5,6,7-tetrahydrofuro[3,2-c]pyridinyl,4,5,6,7-tetrahydrothieno[3,2-b]pyridinyl,1,2,3,4-tetrahydro-1,6-naphthyridinyl, and the like.

The term “aryl” refers to a radical of a monocyclic or polycyclic (e.g.,bicyclic or tricyclic) 4n+2 aromatic ring system (e.g., having 6, 10, or14 π electrons shared in a cyclic array) having 6-14 ring carbon atomsand zero heteroatoms provided in the aromatic ring system (“C₆₋₁₄aryl”). In some embodiments, an aryl group has 6 ring carbon atoms (“C₆aryl”; e.g., phenyl). In some embodiments, an aryl group has 10 ringcarbon atoms (“C₁₀ aryl”; e.g., naphthyl such as 1-naphthyl and2-naphthyl). In some embodiments, an aryl group has 14 ring carbon atoms(“C₁₄ aryl”; e.g., anthracyl). “Aryl” also includes ring systems whereinthe aryl ring, as defined above, is fused with one or more carbocyclylor heterocyclyl groups wherein the radical or point of attachment is onthe aryl ring, and in such instances, the number of carbon atomscontinue to designate the number of carbon atoms in the aryl ringsystem. Unless otherwise specified, each instance of an aryl group isindependently unsubstituted (an “unsubstituted aryl”) or substituted (a“substituted aryl”) with one or more substituents. In certainembodiments, the aryl group is an unsubstituted C₆₋₁₄ aryl. In certainembodiments, the aryl group is a substituted C₆₋₁₄ aryl.

“Aralkyl” is a subset of “alkyl” and refers to an alkyl groupsubstituted by an aryl group, wherein the point of attachment is on thealkyl moiety.

The term “heteroaryl” refers to a radical of a 5-14-membered monocyclicor polycyclic (e.g., bicyclic, tricyclic) 4n+2 aromatic ring system(e.g., having 6, 10, or 14 π electrons shared in a cyclic array) havingring carbon atoms and 1-4 ring heteroatoms provided in the aromatic ringsystem, wherein each heteroatom is independently selected from nitrogen,oxygen, and sulfur (“5-14-membered heteroaryl”). In heteroaryl groupsthat contain one or more nitrogen atoms, the point of attachment can bea carbon or nitrogen atom, as valency permits. Heteroaryl polycyclicring systems can include one or more heteroatoms in one or both rings.“Heteroaryl” includes ring systems wherein the heteroaryl ring, asdefined above, is fused with one or more carbocyclyl or heterocyclylgroups wherein the point of attachment is on the heteroaryl ring, and insuch instances, the number of ring members continue to designate thenumber of ring members in the heteroaryl ring system. “Heteroaryl” alsoincludes ring systems wherein the heteroaryl ring, as defined above, isfused with one or more aryl groups wherein the point of attachment iseither on the aryl or heteroaryl ring, and in such instances, the numberof ring members designates the number of ring members in the fusedpolycyclic (aryl/heteroaryl) ring system. Polycyclic heteroaryl groupswherein one ring does not contain a heteroatom (e.g., indolyl,quinolinyl, carbazolyl, and the like) the point of attachment can be oneither ring, i.e., either the ring bearing a heteroatom (e.g.,2-indolyl) or the ring that does not contain a heteroatom (e.g.,5-indolyl).

In some embodiments, a heteroaryl group is a 5-10-membered aromatic ringsystem having ring carbon atoms and 1-4 ring heteroatoms provided in thearomatic ring system, wherein each heteroatom is independently selectedfrom nitrogen, oxygen, and sulfur (“5-10 membered heteroaryl”). In someembodiments, a heteroaryl group is a 5-8-membered aromatic ring systemhaving ring carbon atoms and 1-4 ring heteroatoms provided in thearomatic ring system, wherein each heteroatom is independently selectedfrom nitrogen, oxygen, and sulfur (“5-8-membered heteroaryl”). In someembodiments, a heteroaryl group is a 5-6-membered aromatic ring systemhaving ring carbon atoms and 1-4 ring heteroatoms provided in thearomatic ring system, wherein each heteroatom is independently selectedfrom nitrogen, oxygen, and sulfur (“5-6-membered heteroaryl”). In someembodiments, the 5-6-membered heteroaryl has 1-3 ring heteroatomsselected from nitrogen, oxygen, and sulfur. In some embodiments, the5-6-membered heteroaryl has 1-2 ring heteroatoms selected from nitrogen,oxygen, and sulfur. In some embodiments, the 5-6-membered heteroaryl has1 ring heteroatom selected from nitrogen, oxygen, and sulfur. Unlessotherwise specified, each instance of a heteroaryl group isindependently unsubstituted (an “unsubstituted heteroaryl”) orsubstituted (a “substituted heteroaryl”) with one or more substituents.In certain embodiments, the heteroaryl group is an unsubstituted 5-14membered heteroaryl. In certain embodiments, the heteroaryl group is asubstituted 5-14 membered heteroaryl.

Exemplary 5-membered heteroaryl groups containing 1 heteroatom include,without limitation, pyrrolyl, furanyl, and thiophenyl. Exemplary5-membered heteroaryl groups containing 2 heteroatoms include, withoutlimitation, imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, thiazolyl, andisothiazolyl. Exemplary 5-membered heteroaryl groups containing 3heteroatoms include, without limitation, triazolyl, oxadiazolyl, andthiadiazolyl. Exemplary 5-membered heteroaryl groups containing 4heteroatoms include, without limitation, tetrazolyl. Exemplary6-membered heteroaryl groups containing 1 heteroatom include, withoutlimitation, pyridinyl. Exemplary 6-membered heteroaryl groups containing2 heteroatoms include, without limitation, pyridazinyl, pyrimidinyl, andpyrazinyl. Exemplary 6-membered heteroaryl groups containing 3 or 4heteroatoms include, without limitation, triazinyl and tetrazinyl,respectively. Exemplary 7-membered heteroaryl groups containing 1heteroatom include, without limitation, azepinyl, oxepinyl, andthiepinyl. Exemplary 5,6-bicyclic heteroaryl groups include, withoutlimitation, indolyl, isoindolyl, indazolyl, benzotriazolyl,benzothiophenyl, isobenzothiophenyl, benzofuranyl, benzoisofuranyl,benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzoxadiazolyl,benzthiazolyl, benzisothiazolyl, benzthiadiazolyl, indolizinyl, andpurinyl. Exemplary 6,6-bicyclic heteroaryl groups include, withoutlimitation, naphthyridinyl, pteridinyl, quinolinyl, isoquinolinyl,cinnolinyl, quinoxalinyl, phthalazinyl, and quinazolinyl. Exemplarytricyclic heteroaryl groups include, without limitation,phenanthridinyl, dibenzofuranyl, carbazolyl, acridinyl, phenothiazinyl,phenoxazinyl and phenazinyl.

“Heteroaralkyl” is a subset of “alkyl” and refers to an alkyl groupsubstituted by a heteroaryl group, wherein the point of attachment is onthe alkyl moiety.

The term “unsaturated bond” refers to a double or triple bond.

The term “unsaturated” or “partially unsaturated” refers to a moietythat includes at least one double or triple bond.

The term “saturated” refers to a moiety that does not contain a doubleor triple bond, i.e., the moiety only contains single bonds.

Affixing the suffix “-ene” to a group indicates the group is a divalentmoiety, e.g., alkylene is the divalent moiety of alkyl, alkenylene isthe divalent moiety of alkenyl, alkynylene is the divalent moiety ofalkynyl, heteroalkylene is the divalent moiety of heteroalkyl,heteroalkenylene is the divalent moiety of heteroalkenyl,heteroalkynylene is the divalent moiety of heteroalkynyl, carbocyclyleneis the divalent moiety of carbocyclyl, heterocyclylene is the divalentmoiety of heterocyclyl, arylene is the divalent moiety of aryl, andheteroarylene is the divalent moiety of heteroaryl.

A group is optionally substituted unless expressly provided otherwise.In certain embodiments, alkyl, alkenyl, alkynyl, heteroalkyl,heteroalkenyl, heteroalkynyl, carbocyclyl, heterocyclyl, aryl, andheteroaryl groups are optionally substituted. “Optionally substituted”refers to a group which may be substituted or unsubstituted (e.g.,“substituted” or “unsubstituted” alkyl, “substituted” or “unsubstituted”alkenyl, “substituted” or “unsubstituted” alkynyl, “substituted” or“unsubstituted” heteroalkyl, “substituted” or “unsubstituted”heteroalkenyl, “substituted” or “unsubstituted” heteroalkynyl,“substituted” or “unsubstituted” carbocyclyl, “substituted” or“unsubstituted” heterocyclyl, “substituted” or “unsubstituted” aryl or“substituted” or “unsubstituted” heteroaryl group). In general, the term“substituted” means that at least one hydrogen present on a group isreplaced with a permissible substituent, e.g., a substituent which uponsubstitution results in a stable compound, e.g., a compound which doesnot spontaneously undergo transformation such as by rearrangement,cyclization, elimination, or other reaction. Unless otherwise indicated,a “substituted” group has a substituent at one or more substitutablepositions of the group, and when more than one position in any givenstructure is substituted, the substituent is either the same ordifferent at each position. The term “substituted” is contemplated toinclude substitution with all permissible substituents of organiccompounds, and includes any of the substituents described herein thatresults in the formation of a stable compound. The present disclosurecontemplates any and all such combinations in order to arrive at astable compound. For purposes of this disclosure, heteroatoms such asnitrogen may have hydrogen substituents and/or any suitable substituentas described herein which satisfy the valencies of the heteroatoms andresults in the formation of a stable moiety.

Exemplary carbon atom substituents include, but are not limited to,halogen, —CN, —NO₂, —N₃, —SO₂H, —SO₃H, —OH, —OR^(aa), —ON(R^(bb))₂,—N(R^(bb))₂, —N(R^(bb))₃ ⁺X⁻, —N(OR^(cc))R^(bb), —SH, —SR^(aa),—SSR^(cc), —C(═O)R^(aa), —CO₂H, —CHO, —C(OR^(cc))₂, —CO₂R^(aa),—OC(═O)R^(aa), —OCO₂R^(aa), —C(═O)N(R^(bb))₂, —OC(═O)N(R^(bb))₂,—NR^(bb)C(═O)R^(aa), —NR^(bb)C₂R^(aa), —NR^(bb)C(═O)N(R^(bb))₂,—C(═NR^(bb))R^(aa), —C(═NR^(bb))OR^(aa), —OC(═NR^(bb))R^(aa),—OC(═NR^(bb))OR^(aa), —C(═NR^(bb))N(R^(bb))₂, —OC(═NR^(bb))N(R^(bb))₂,—NR^(bb)C(═NR^(bb))N(R^(bb))₂, —C(═O)NR^(bb)SO₂R^(aa),—NR^(bb)SO₂R^(aa), —SO₂N(R^(bb))₂, —SO₂R^(aa), —SO₂OR^(aa), —OSO₂R^(aa),—S(═O)R^(aa), —OS(═O)R^(aa), —Si(R^(aa))₃,—OSi(R^(aa))₃—C(═S)N(R^(bb))₂, —C(═O)SR^(aa), —C(═S)SR^(aa),—SC(═S)SR^(aa), —SC(═O)SR^(aa), —OC(═O)SR^(aa), —SC(═O)OR^(aa),—SC(═O)R^(aa), —P(═O)(R^(aa))₂, —P(═O)(OR^(cc))₂, —OP(═O)(R^(aa))₂,—OP(═O)(OR^(cc))₂, —P(═O)(N(R^(bb))₂)₂, —OP(═O)(N(R^(bb))₂)₂,—NR^(bb)P(═O)(R^(aa))₂, —NR^(bb)P(═)(OR^(cc))₂,—NR^(bb)P(═O)(N(R^(bb))₂)₂, —P(R^(cc))₂, —P(OR^(cc))₂, —P(R^(cc))₃ ⁺X⁻,—P(OR^(cc))₃ ⁺X⁻, —P(R^(cc))₄, —P(OR^(cc))₄, —OP(R^(cc))₂, —OP(R^(cc))₃⁺X⁻, —OP(OR^(cc))₂, —OP(OR^(cc))₃ ⁺X⁻, —OP(R^(cc))₄, —OP(OR^(cc))₄,—B(R^(aa))₂, —B(OR^(cc))₂, —BR^(aa)(OR^(cc)), C₁₋₁₀ alkyl, C₁₋₁₀perhaloalkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, heteroC₁₋₁₀ alkyl,heteroC₂₋₁₀ alkenyl, heteroC₂₋₁₀ alkynyl, C₃₋₁₀ carbocyclyl, 3-14membered heterocyclyl, C₆₋₁₄ aryl, and 5-14 membered heteroaryl, whereineach alkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl,carbocyclyl, heterocyclyl, aryl, and heteroaryl is independentlysubstituted with 0, 1, 2, 3, 4, or 5 R^(dd) groups; wherein X⁻ is acounterion;

or two geminal hydrogens on a carbon atom are replaced with the group═O, ═S, ═NN(R^(bb))₂, ═NNR^(bb)C(═O)R^(aa), ═NNR^(bb)C(═O)OR^(aa),═NNR^(bb)S(═O)₂R^(aa), ═NR^(bb), or ═NOR^(cc);

each instance of R^(aa) is, independently, selected from C₁₋₁₀ alkyl,C₁₋₁₀ perhaloalkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, heteroC₁ 10 alkyl,heteroC₂₋₁₀ alkenyl, heteroC₂₋₁₀ alkynyl, C₃₋₁₀ carbocyclyl, 3-14membered heterocyclyl, C₆₋₁₄ aryl, and 5-14 membered heteroaryl, or twoR^(aa) groups are joined to form a 3-14 membered heterocyclyl or 5-14membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl,heteroalkyl, heteroalkenyl, heteroalkynyl, carbocyclyl, heterocyclyl,aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or5 R^(dd) groups;

each instance of R^(bb) is, independently, selected from hydrogen, —OH,—OR^(aa), —N(R^(cc))₂, —CN, —C(═O)R^(aa), —C(═O)N(R^(cc))₂, —CO₂R,—SO₂R^(aa), —C(═NR^(cc))OR^(aa), —C(═NR^(cc))N(R^(cc))₂, —SO₂N(R^(cc))₂,—SO₂R^(cc), —SO₂OR^(cc), —SOR^(aa), —C(═S)N(R^(cc))₂, —C(═O)SR^(cc),—C(═S)SR^(cc), —P(═O)(R^(aa))₂, —P(═O)(OR^(cc))₂, —P(═O)(N(R^(cc))₂)₂,C₁₋₁₀ alkyl, C₁₋₁₀ perhaloalkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl,heteroC₁₋₁₀alkyl, heteroC₂₋₁₀alkenyl, heteroC₂₋₁₀alkynyl, C₃₋₁₀carbocyclyl, 3-14 membered heterocyclyl, C₆₋₁₄ aryl, and 5-14 memberedheteroaryl, or two R^(bb) groups are joined to form a 3-14 memberedheterocyclyl or 5-14 membered heteroaryl ring, wherein each alkyl,alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl,carbocyclyl, heterocyclyl, aryl, and heteroaryl is independentlysubstituted with 0, 1, 2, 3, 4, or 5 R^(dd) groups; wherein X⁻ is acounterion;

each instance of R^(cc) is, independently, selected from hydrogen, C₁₋₁₀alkyl, C₁₋₁₀ perhaloalkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, heteroC₁₋₁₀alkyl, heteroC₂₋₁₀ alkenyl, heteroC₂₋₁₀ alkynyl, C₃₋₁₀ carbocyclyl, 3-14membered heterocyclyl, C₆₋₁₄ aryl, and 5-14 membered heteroaryl, or twoR^(cc) groups are joined to form a 3-14 membered heterocyclyl or 5-14membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl,heteroalkyl, heteroalkenyl, heteroalkynyl, carbocyclyl, heterocyclyl,aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or5 R^(dd) groups;

each instance of R^(dd) is, independently, selected from halogen, —CN,—NO₂, —N₃, —SO₂H, —SO₃H, —OH, —OR^(ee), —ON(R^(ff))₂, —N(R^(ff))₂,—N(R^(ff))₃ ⁺X⁻, —N(OR^(ee))R^(ff), —SH, —SR^(ee), —SSR^(ee),—C(═O)R^(ee), —C, —CO₂H, —CO₂R^(ee), —OC(═O)R^(ee), —OCO₂R^(ee),—C(═O)N(R^(ff))₂, —OC(═O)N(R^(ff))₂, —NR^(ff)C(═O)R^(ee),—NR^(ff)CO₂R^(ee), —NR^(ff)C(═O)N(R^(ff))₂, —C(═NR^(ff))OR^(ee),—OC(═NR^(ff))R^(ee), —OC(═NR^(ff))OR^(ee), —C(═NR^(ff))N(R^(ff))₂,—OC(═NR^(ff))N(R^(ff))₂, —NR^(ff)C(═NR^(ff))N(R^(ff))₂,—NR^(ff)SO₂R^(ee), —SO₂N(R^(ff))₂, —SO₂R^(ee), —SO₂OR^(ee), —OSO₂R^(ee),—S(═O)R^(ee), —Si(R^(ee))₃, —OSi(R^(ee))₃, —C(═S)N(R^(ff))₂,—C(═O)SR^(ee), —C(═S)SR^(ee), —SC(═S)SR^(ee), —P(═O)(OR^(ee))₂,—P(═O)(R^(ee))₂, —OP(═O)(R^(ee))₂, —OP(═O)(OR^(ee))₂, C₁₋₆ alkyl, C₁₋₆perhaloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, heteroC₁₋₆alkyl,heteroC₂₋₆alkenyl, heteroC₂₋₆alkynyl, C₃₋₁₀ carbocyclyl, 3-10 memberedheterocyclyl, C₆₋₁₀ aryl, 5-10 membered heteroaryl, wherein each alkyl,alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl,carbocyclyl, heterocyclyl, aryl, and heteroaryl is independentlysubstituted with 0, 1, 2, 3, 4, or 5 R^(gg) groups, or two geminalR^(dd) substituents can be joined to form ═O or ═S; wherein X⁻ is acounterion;

each instance of R^(ee) is, independently, selected from C₁₋₆ alkyl,C₁₋₆ perhaloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, heteroC₁₋₆ alkyl,heteroC₂₋₆alkenyl, heteroC₂₋₆ alkynyl, C₃₋₁₀ carbocyclyl, C₆₋₁₀ aryl,3-10 membered heterocyclyl, and 3-10 membered heteroaryl, wherein eachalkyl, alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl,carbocyclyl, heterocyclyl, aryl, and heteroaryl is independentlysubstituted with 0, 1, 2, 3, 4, or 5 R^(gg) groups;

each instance of R^(ff) is, independently, selected from hydrogen, C₁₋₆alkyl, C₁₋₆ perhaloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, heteroC₁₋₆alkyl,heteroC₂₋₆alkenyl, heteroC₂₋₆alkynyl, C₃₋₁₀ carbocyclyl, 3-10 memberedheterocyclyl, C₆₋₁₀ aryl and 5-10 membered heteroaryl, or two R^(ff)groups are joined to form a 3-10 membered heterocyclyl or 5-10 memberedheteroaryl ring, wherein each alkyl, alkenyl, alkynyl, heteroalkyl,heteroalkenyl, heteroalkynyl, carbocyclyl, heterocyclyl, aryl, andheteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 R^(gg)groups; and

each instance of R^(gg) is, independently, halogen, —CN, —NO₂, —N₃,—SO₂H, —SO₃H, —OH, —OC₁₋₆ alkyl, —ON(C₁₋₆ alkyl)₂, —N(C₁₋₆ alkyl)₂,—N(C₁₋₆ alkyl)₃+X⁻, —NH(C₁₋₆ alkyl)₂+X⁻, —NH₂(C₁₋₆ alkyl)⁺X⁻, —NH₃+X⁻,—N(OC₁₋₆ alkyl)(C₁₋₆ alkyl), —N(OH)(C₁₋₆ alkyl), —NH(OH), —SH, —SC₁₋₆alkyl, —SS(C₁₋₆ alkyl), —C(═O)(C₁₋₆ alkyl), —CO₂H, —CO₂(C₁₋₆ alkyl),—OC(═O)(C₁₋₆ alkyl), —OCO₂(C₁₋₆ alkyl), —C(═O)NH₂, —C(═O)N(C₁₋₆ alkyl)₂,—OC(═O)NH(C₁₋₆ alkyl), —NHC(═O)(C₁₋₆ alkyl), —N(C₁₋₆ alkyl)C(═O)(C₁₋₆alkyl), —NHCO₂(C₁₋₆ alkyl), —NHC(═O)N(C₁₋₆ alkyl)₂, —NHC(═O)NH(C₁₋₆alkyl), —NHC(═O)NH₂, —C(═NH)O(C₁₋₆ alkyl), —OC(═NH)(C₁₋₆ alkyl),—OC(═NH)OC₁₋₆ alkyl, —C(═NH)N(C₁₋₆ alkyl)₂, —C(═NH)NH(C₁₋₆ alkyl),—C(═NH)NH₂, —OC(═NH)N(C₁₋₆ alkyl)₂, —OC(NH)NH(C₁₋₆ alkyl), —OC(NH)NH₂,—NHC(NH)N(C₁₋₆ alkyl)₂, —NHC(═NH)NH₂, —NHSO₂(C₁₋₆ alkyl), —SO₂N(C₁₋₆alkyl)₂, —SO₂NH(C₁₋₆ alkyl), —SO₂NH₂, —SO₂C₁₋₆ alkyl, —SO₂OC₁₋₆ alkyl,—OSO₂C₁₋₆ alkyl, —SOC₁₋₆ alkyl, —Si(C₁₋₆ alkyl)₃, —OSi(C₁₋₆alkyl)₃-C(═S)N(C₁₋₆ alkyl)₂, C(═S)NH(C₁₋₆ alkyl), C(═S)NH₂, —C(═O)S(C₁₋₆alkyl), —C(═S)SC₁₋₆ alkyl, —SC(═S)SC₁₋₆ alkyl, —P(═O)(OC₁₋₆ alkyl)₂,—P(═O)(C₁₋₆ alkyl)₂, —OP(═O)(C₁₋₆ alkyl)₂, —OP(═O)(OC₁₋₆ alkyl)₂, C₁₋₆alkyl, C₁₋₆ perhaloalkyl, C₂₋₆ alkenyl, C₂₋₆ alkynyl, heteroC₁₋₆alkyl,heteroC₂₋₆alkenyl, heteroC₂₋₆alkynyl, C₃₋₁₀ carbocyclyl, C₆₋₁₀ aryl,3-10 membered heterocyclyl, 5-10 membered heteroaryl; or two geminalR^(gg) substituents can be joined to form ═O or ═S; wherein X⁻ is acounterion.

The term “halo” or “halogen” refers to fluorine (fluoro, —F), chlorine(chloro, —Cl), bromine (bromo, —Br), or iodine (iodo, —I).

The term “hydroxyl” or “hydroxy” refers to the group —OH. The term“substituted hydroxyl” or “substituted hydroxyl,” by extension, refersto a hydroxyl group wherein the oxygen atom directly attached to theparent molecule is substituted with a group other than hydrogen, andincludes groups selected from —OR^(aa), —ON(R^(bb))₂, —OC(═O)SR^(aa),—OC(═O)R^(aa), —OCO₂R^(aa), —OC(═O)N(R^(bb))₂, —OC(═NR^(bb))R^(aa),—OC(═NR^(bb))OR^(aa), —OC(═NR^(bb))N(R^(bb))₂, —OS(═O)R^(aa),—OSO₂R^(aa), —OSi(R^(aa))₃, —OP(R^(cc))₂, —OP(R^(cc))₃ ⁺X⁻,—OP(OR^(cc))₂, —OP(OR^(cc))₃ ⁺X⁻, —OP(═O)(R^(aa))₂, —OP(═O)(OR^(cc))₂,and —OP(═O)(N(R^(bb)))₂, wherein X⁻, R^(aa), R^(bb), and R^(cc) are asdefined herein.

The term “thiol” or “thio” refers to the group —SH. The term“substituted thiol” or “substituted thio,” by extension, refers to athiol group wherein the sulfur atom directly attached to the parentmolecule is substituted with a group other than hydrogen, and includesgroups selected from —SR^(aa), —S═SR^(cc), —SC(═S)SR^(aa),—SC(═O)SR^(aa), —SC(═O)OR^(aa), and —SC(═O)R^(aa), wherein R^(aa) andR^(cc) are as defined herein.

The term “amino” refers to the group —NH₂. The term “substituted amino,”by extension, refers to a monosubstituted amino, a disubstituted amino,or a trisubstituted amino.

In certain embodiments, the “substituted amino” is a monosubstitutedamino or a disubstituted amino group.

The term “monosubstituted amino” refers to an amino group wherein thenitrogen atom directly attached to the parent molecule is substitutedwith one hydrogen and one group other than hydrogen, and includes groupsselected from —NH(R^(bb)), —NHC(═O)R^(aa), —NHCO₂R^(aa),—NHC(═O)N(R^(bb))₂, —NHC(═NR^(bb))N(R^(bb))₂, —NHSO₂R^(aa),—NHP(═O)(OR^(cc))₂, and —NHP(═O)(N(R^(bb))₂)₂, wherein R^(aa), R^(bb)and R^(cc) are as defined herein, and wherein R^(bb) of the group—NH(R^(bb)) is not hydrogen.

The term “disubstituted amino” refers to an amino group wherein thenitrogen atom directly attached to the parent molecule is substitutedwith two groups other than hydrogen, and includes groups selected from—N(R^(bb))₂, —NR^(bb) C(═O)R^(aa), —NR^(bb)CO₂R^(aa),—NR^(bb)C(═O)N(R^(bb))₂, —NR^(bb)C(═NR^(bb))N(R^(bb))₂,—NR^(bb)SO₂R^(aa), —NR^(bb)P(═O)(OR^(cc))₂, and—NR^(bb)P(═O)(N(R^(bb))₂)₂, wherein R^(aa), R^(bb), and R^(cc) are asdefined herein, with the proviso that the nitrogen atom directlyattached to the parent molecule is not substituted with hydrogen.

The term “trisubstituted amino” refers to an amino group wherein thenitrogen atom directly attached to the parent molecule is substitutedwith three groups, and includes groups selected from —N(R^(bb))₃ and—N(R^(bb))₃ ⁺X⁻, wherein R^(bb) and X⁻ are as defined herein.

The term “carbonyl” refers a group wherein the carbon directly attachedto the parent molecule is sp² hybridized, and is substituted with anoxygen, nitrogen or sulfur atom, e.g., a group selected from ketones(—C(═O)R^(aa)), carboxylic acids (—CO₂H), aldehydes (—CHO), esters(—CO₂R^(aa), —C(═O)SR^(aa), —C(═S)SR^(aa)), amides (—C(═O)N(R^(bb))₂,—C(═O)NR^(bb)SO₂R^(aa), —C(═S)N(R^(bb))₂), and imines(—C(═NR^(bb))R^(aa), —C(═NR^(bb))OR^(aa)), —C(═NR^(bb))N(R^(bb))₂),wherein R^(aa) and R^(bb) are as defined herein.

The term “silyl” refers to the group —Si(R^(aa))₃, wherein R^(aa) is asdefined herein.

The term “oxo” refers to the group ═O, and the term “thiooxo” refers tothe group ═S.

Nitrogen atoms can be substituted or unsubstituted as valency permits,and include primary, secondary, tertiary, and quaternary nitrogen atoms.Exemplary nitrogen atom substituents include, but are not limited to,hydrogen, —OH, —OR^(aa), —N(R^(cc))₂, —CN, —C(═O)R^(aa),—C(═O)N(R^(cc))₂, —CO₂R^(aa), —SO₂R^(aa), —C(═NR^(bb))R^(a),—C(═NR^(cc))OR^(aa), —C(═NR^(cc))N(R^(cc))₂, —SO₂N(R^(cc))₂, —SO₂R^(cc),—SO₂OR^(cc), —SOR^(aa), —C(═S)N(R^(cc))₂, —C(═O)SR^(cc), —C(═S)SR^(cc),—P(═O)(OR^(cc))₂, —P(═O)(R^(aa))₂, —P(═O)(N(R^(cc))₂)₂, C₁₋₁₀ alkyl,C₁₋₁₀ perhaloalkyl, C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl, heteroC₁₋₁₀-alkyl,heteroC₂₋₁₀alkenyl, heteroC₂₋₁₀alkynyl, C₃₋₁₀ carbocyclyl, 3-14 memberedheterocyclyl, C₆₋₁₄ aryl, and 5-14 membered heteroaryl, or two R^(cc)groups attached to an N atom are joined to form a 3-14 memberedheterocyclyl or 5-14 membered heteroaryl ring, wherein each alkyl,alkenyl, alkynyl, heteroalkyl, heteroalkenyl, heteroalkynyl,carbocyclyl, heterocyclyl, aryl, and heteroaryl is independentlysubstituted with 0, 1, 2, 3, 4, or 5 R^(dd) groups, and wherein R^(aa),R^(bb), R^(cc) and R^(dd) are as defined above.

In certain embodiments, the substituent present on the nitrogen atom isan nitrogen protecting group (also referred to herein as an “aminoprotecting group”). Nitrogen protecting groups include, but are notlimited to, —OH, —OR^(aa), —N(R^(cc))₂, —C(═O)R^(aa), —C(═O)N(R^(cc))₂,—CO₂R^(aa), —SO₂R^(aa), —C(═NR^(cc))R^(aa), —C(═NR^(cc))OR^(aa),—C(═NR^(cc))N(R^(cc))₂, —SO₂N(R^(cc))₂, —SO₂R^(cc), —SO₂OR^(cc),—SOR^(aa), —C(═S)N(R^(cc))₂, —C(═O)SR^(cc), —C(═S)SR^(cc), C₁₋₁₀ alkyl(e.g., aralkyl, heteroaralkyl), C₂₋₁₀ alkenyl, C₂₋₁₀ alkynyl,heteroC₁₋₁₀ alkyl, heteroC₂₋₁₀ alkenyl, heteroC₂₋₁₀ alkynyl, C₃₋₁₀carbocyclyl, 3-14 membered heterocyclyl, C₆₋₁₄ aryl, and 5-14 memberedheteroaryl groups, wherein each alkyl, alkenyl, alkynyl, heteroalkyl,heteroalkenyl, heteroalkynyl, carbocyclyl, heterocyclyl, aralkyl, aryl,and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5R^(dd) groups, and wherein R^(aa), R^(bb), R^(cc) and R^(dd) are asdefined herein. Nitrogen protecting groups are well known in the art andinclude those described in detail in Protecting Groups in OrganicSynthesis, T. W. Greene and P. G. M. Wuts, 3^(rd) edition, John Wiley &Sons, 1999, incorporated herein by reference.

For example, nitrogen protecting groups such as amide groups (e.g.,—C(═O)R^(aa)) include, but are not limited to, formamide, acetamide,chloroacetamide, trichloroacetamide, trifluoroacetamide,phenylacetamide, 3-phenylpropanamide, picolinamide,3-pyridylcarboxamide, N-benzoylphenylalanyl derivative, benzamide,p-phenylbenzamide, o-nitophenylacetamide, o-nitrophenoxyacetamide,acetoacetamide, (N′-dithiobenzyloxyacylamino)acetamide,3-(p-hydroxyphenyl)propanamide, 3-(o-nitrophenyl)propanamide,2-methyl-2-(o-nitrophenoxy)propanamide,2-methyl-2-(o-phenylazophenoxy)propanamide, 4-chlorobutanamide,3-methyl-3-nitrobutanamide, o-nitrocinnamide, N-acetylmethioninederivative, o-nitrobenzamide and o-(benzoyloxymethyl)benzamide.

Nitrogen protecting groups such as carbamate groups (e.g.,—C(═O)OR^(aa)) include, but are not limited to, methyl carbamate, ethylcarbamate, 9-fluorenylmethyl carbamate (Fmoc),9-(2-sulfo)fluorenylmethyl carbamate, 9-(2,7-dibromo)fluoroenylmethylcarbamate,2,7-di-t-butyl-[9-(10,10-dioxo-10,10,10,10-tetrahydrothioxanthyl)]methylcarbamate (DBD-Tmoc), 4-methoxyphenacyl carbamate (Phenoc),2,2,2-trichloroethyl carbamate (Troc), 2-trimethylsilylethyl carbamate(Teoc), 2-phenylethyl carbamate (hZ), 1-(1-adamantyl)-1-methylethylcarbamate (Adpoc), 1,1-dimethyl-2-haloethyl carbamate,1,1-dimethyl-2,2-dibromoethyl carbamate (DB-t-BOC),1,1-dimethyl-2,2,2-trichloroethyl carbamate (TCBOC),1-methyl-1-(4-biphenylyl)ethyl carbamate (Bpoc),1-(3,5-di-t-butylphenyl)-1-methylethyl carbamate (t-Bumeoc), 2-(2′- and4′-pyridyl)ethyl carbamate (Pyoc), 2-(N,N-dicyclohexylcarboxamido)ethylcarbamate, t-butyl carbamate (BOC or Boc), 1-adamantyl carbamate (Adoc),vinyl carbamate (Voc), allyl carbamate (Alloc), 1-isopropylallylcarbamate (Ipaoc), cinnamyl carbamate (Coc), 4-nitrocinnamyl carbamate(Noc), 8-quinolyl carbamate, N-hydroxypiperidinyl carbamate, alkyldithiocarbamate, benzyl carbamate (Cbz), p-methoxybenzyl carbamate (Moz),p-nitobenzyl carbamate, p-bromobenzyl carbamate, p-chlorobenzylcarbamate, 2,4-dichlorobenzyl carbamate, 4-methylsulfinylbenzylcarbamate (Msz), 9-anthrylmethyl carbamate, diphenylmethyl carbamate,2-methylthioethyl carbamate, 2-methylsulfonylethyl carbamate,2-(p-toluenesulfonyl)ethyl carbamate, [2-(1,3-dithianyl)]methylcarbamate (Dmoc), 4-methylthiophenyl carbamate (Mtpc),2,4-dimethylthiophenyl carbamate (Bmpc), 2-phosphonioethyl carbamate(Peoc), 2-triphenylphosphonioisopropyl carbamate (Ppoc),1,1-dimethyl-2-cyanoethyl carbamate, m-chloro-p-acyloxybenzyl carbamate,p-(dihydroxyboryl)benzyl carbamate, 5-benzisoxazolylmethyl carbamate,2-(trifluoromethyl)-6-chromonylmethyl carbamate (Tcroc), m-nitrophenylcarbamate, 3,5-dimethoxybenzyl carbamate, o-nitrobenzyl carbamate,3,4-dimethoxy-6-nitrobenzyl carbamate, phenyl(o-nitrophenyl)methylcarbamate, t-amyl carbamate, S-benzyl thiocarbamate, p-cyanobenzylcarbamate, cyclobutyl carbamate, cyclohexyl carbamate, cyclopentylcarbamate, cyclopropylmethyl carbamate, p-decyloxybenzyl carbamate,2,2-dimethoxyacylvinyl carbamate, o-(N,N-dimethylcarboxamido)benzylcarbamate, 1,1-dimethyl-3-(N,N-dimethylcarboxamido)propyl carbamate,1,1-dimethylpropynyl carbamate, di(2-pyridyl)methyl carbamate,2-furanylmethyl carbamate, 2-iodoethyl carbamate, isoborynl carbamate,isobutyl carbamate, isonicotinyl carbamate,p-(p′-methoxyphenylazo)benzyl carbamate, 1-methylcyclobutyl carbamate,1-methylcyclohexyl carbamate, 1-methyl-1-cyclopropylmethyl carbamate,1-methyl-1-(3,5-dimethoxyphenyl)ethyl carbamate,1-methyl-1-(p-phenylazophenyl)ethyl carbamate, 1-methyl-1-phenylethylcarbamate, 1-methyl-1-(4-pyridyl)ethyl carbamate, phenyl carbamate,p-(phenylazo)benzyl carbamate, 2,4,6-tri-t-butylphenyl carbamate,4-(trimethylammonium)benzyl carbamate, and 2,4,6-trimethylbenzylcarbamate.

Nitrogen protecting groups such as sulfonamide groups (e.g.,—S(═O)₂R^(aa)) include, but are not limited to, p-toluenesulfonamide(Ts), benzenesulfonamide, 2,3,6,-trimethyl-4-methoxybenzenesulfonamide(Mtr), 2,4,6-trimethoxybenzenesulfonamide (Mtb),2,6-dimethyl-4-methoxybenzenesulfonamide (Pme),2,3,5,6-tetramethyl-4-methoxybenzenesulfonamide (Mte),4-methoxybenzenesulfonamide (Mbs), 2,4,6-trimethylbenzenesulfonamide(Mts), 2,6-dimethoxy-4-methylbenzenesulfonamide (iMds),2,2,5,7,8-pentamethylchroman-6-sulfonamide (Pmc), methanesulfonamide(Ms), β-trimethylsilylethanesulfonamide (SES), 9-anthracenesulfonamide,4-(4′,8′-dimethoxynaphthylmethyl)benzenesulfonamide (DNMBS),benzylsulfonamide, trifluoromethylsulfonamide, and phenacylsulfonamide.

Other nitrogen protecting groups include, but are not limited to,phenothiazinyl-(10)-acyl derivative, N′-p-toluenesulfonylaminoacylderivative, N′-phenylaminothioacyl derivative, N-benzoylphenylalanylderivative, N-acetylmethionine derivative,4,5-diphenyl-3-oxazolin-2-one, N-phthalimide, N-dithiasuccinimide (Dts),N-2,3-diphenylmaleimide, N-2,5-dimethylpyrrole,N-1,1,4,4-tetramethyldisilylazacyclopentane adduct (STABASE),5-substituted 1,3-dimethyl-1,3,5-triazacyclohexan-2-one, 5-substituted1,3-dibenzyl-1,3,5-triazacyclohexan-2-one, 1-substituted3,5-dinitro-4-pyridone, N-methylamine, N-allylamine,N-[2-(trimethylsilyl)ethoxy]methylamine (SEM), N-3-acetoxypropylamine,N-(1-isopropyl-4-nitro-2-oxo-3-pyroolin-3-yl)amine, quaternary ammoniumsalts, N-benzylamine, N-di(4-methoxyphenyl)methylamine,N-5-dibenzosuberylamine, N-triphenylmethylamine (Tr),N-[(4-methoxyphenyl)diphenylmethyl]amine (MMTr),N-9-phenylfluorenylamine (PhF),N-2,7-dichloro-9-fluorenylmethyleneamine, N-ferrocenylmethylamino (Fcm),N-2-picolylamino N′-oxide, N-1,1-dimethylthiomethyleneamine,N-benzylideneamine, N-p-methoxybenzylideneamine,N-diphenylmethyleneamine, N-[(2-pyridyl)mesityl]methyleneamine,N—(N′,N′-dimethylaminomethylene)amine, N,N′-isopropylidenediamine,N-p-nitrobenzylideneamine, N-salicylideneamine,N-5-chlorosalicylideneamine,N-(5-chloro-2-hydroxyphenyl)phenylmethyleneamine,N-cyclohexylideneamine, N-(5,5-dimethyl-3-oxo-1-cyclohexenyl)amine,N-borane derivative, N-diphenylborinic acid derivative,N-[phenyl(pentaacylchromium- or tungsten)acyl]amine, N-copper chelate,N-zinc chelate, N-nitroamine, N-nitrosoamine, amine N-oxide,diphenylphosphinamide (Dpp), dimethylthiophosphinamide (Mpt),diphenylthiophosphinamide (Ppt), dialkyl phosphoramidates, dibenzylphosphoramidate, diphenyl phosphoramidate, benzenesulfenamide,o-nitrobenzenesulfenamide (Nps), 2,4-dinitrobenzenesulfenamide,pentachlorobenzenesulfenamide, 2-nitro-4-methoxybenzenesulfenamide,triphenylmethylsulfenamide, and 3-nitropyridinesulfenamide (Npys).

In certain embodiments, the substituent present on an oxygen atom is anoxygen protecting group (also referred to herein as an “hydroxylprotecting group”). Oxygen protecting groups include, but are notlimited to, —R^(aa), —N(R^(bb))₂, —C(═O)SR^(aa), —C(═O)R^(aa),—CO₂R^(aa), —C(═O)N(R^(bb))₂, —C(═NR^(bb))R^(aa), —C(═NR^(bb))OR^(aa),—C(═NR^(bb))N(R^(bb))₂, —S(═O)R^(aa), —SO₂R^(aa), —Si(R^(aa))₃,—P(R^(cc))₂, —P(R^(cc))₃ ⁺X⁻, —P(ORc)₂, —P(OR^(cc))₃ ⁺X⁻,—P(═O)(R^(aa))₂, —P(═O)(OR^(cc))₂, and —P(═O)(N(R^(bb))₂)₂, wherein X⁻,R^(aa), R^(bb), and R^(cc) are as defined herein. Oxygen protectinggroups are well known in the art and include those described in detailin Protecting Groups in Organic Synthesis, T. W. Greene and P. G. M.Wuts, 3^(rd) edition, John Wiley & Sons, 1999, incorporated herein byreference.

Exemplary oxygen protecting groups include, but are not limited to,methyl, methoxylmethyl (MOM), methylthiomethyl (MTM), t-butylthiomethyl,(phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM),p-methoxybenzyloxymethyl (PMBM), (4-methoxyphenoxy)methyl (p-AOM),guaiacolmethyl (GUM), t-butoxymethyl, 4-pentenyloxymethyl (POM),siloxymethyl, 2-methoxyethoxymethyl (MEM), 2,2,2-trichloroethoxymethyl,bis(2-chloroethoxy)methyl, 2-(trimethylsilyl)ethoxymethyl (SEMOR),tetrahydropyranyl (THP), 3-bromotetrahydropyranyl,tetrahydrothiopyranyl, 1-methoxycyclohexyl, 4-methoxytetrahydropyranyl(MTHP), 4-methoxytetrahydrothiopyranyl, 4-methoxytetrahydrothiopyranylS,S-dioxide, 1-[(2-chloro-4-methyl)phenyl]-4-methoxypiperidin-4-yl(CTMP), 1,4-dioxan-2-yl, tetrahydrofuranyl, tetrahydrothiofuranyl,2,3,3a,4,5,6,7,7a-octahydro-7,8,8-trimethyl-4,7-methanobenzofuran-2-yl,1-ethoxyethyl, 1-(2-chloroethoxy)ethyl, 1-methyl-1-methoxyethyl,1-methyl-1-benzyloxyethyl, 1-methyl-1-benzyloxy-2-fluoroethyl,2,2,2-trichloroethyl, 2-trimethylsilylethyl, 2-(phenylselenyl)ethyl,t-butyl, allyl, p-chlorophenyl, p-methoxyphenyl, 2,4-dinitrophenyl,benzyl (Bn), p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl,p-nitrobenzyl, p-halobenzyl, 2,6-dichlorobenzyl, p-cyanobenzyl,p-phenylbenzyl, 2-picolyl, 4-picolyl, 3-methyl-2-picolyl N-oxido,diphenylmethyl, p,p′-dinitrobenzhydryl, 5-dibenzosuberyl,triphenylmethyl, α-naphthyldiphenylmethyl,p-methoxyphenyldiphenylmethyl, di(p-methoxyphenyl)phenylmethyl,tri(p-methoxyphenyl)methyl, 4-(4′-bromophenacyloxyphenyl)diphenylmethyl,4,4′,4″-tris(4,5-dichlorophthalimidophenyl)methyl,4,4′,4″-tris(levulinoyloxyphenyl)methyl,4,4′,4″-tris(benzoyloxyphenyl)methyl,3-(imidazol-1-yl)bis(4′,4″-dimethoxyphenyl)methyl,1,1-bis(4-methoxyphenyl)-1′-pyrenylmethyl, 9-anthryl,9-(9-phenyl)xanthenyl, 9-(9-phenyl-10-oxo)anthryl,1,3-benzodithiolan-2-yl, benzisothiazolyl S,S-dioxido, trimethylsilyl(TMS), triethylsilyl (TES), triisopropylsilyl (TIPS),dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS),dimethylthexylsilyl, t-butyldimethylsilyl (TBDMS), t-butyldiphenylsilyl(TBDPS), tribenzylsilyl, tri-p-xylylsilyl, triphenylsilyl,diphenylmethylsilyl (DPMS), t-butylmethoxyphenylsilyl (TBMPS), formate,benzoylformate, acetate, chloroacetate, dichloroacetate,trichloroacetate, trifluoroacetate, methoxyacetate,triphenylmethoxyacetate, phenoxyacetate, p-chlorophenoxyacetate,3-phenylpropionate, 4-oxopentanoate (levulinate),4,4-(ethylenedithio)pentanoate (levulinoyldithioacetal), pivaloate,adamantoate, crotonate, 4-methoxycrotonate, benzoate, p-phenylbenzoate,2,4,6-trimethylbenzoate (mesitoate), methyl carbonate, 9-fluorenylmethylcarbonate (Fmoc), ethyl carbonate, 2,2,2-trichloroethyl carbonate(Troc), 2-(trimethylsilyl)ethyl carbonate (TMSEC), 2-(phenylsulfonyl)ethyl carbonate (Psec), 2-(triphenylphosphonio) ethyl carbonate (Peoc),isobutyl carbonate, vinyl carbonate, allyl carbonate, t-butyl carbonate(BOC or Boc), p-nitrophenyl carbonate, benzyl carbonate, p-methoxybenzylcarbonate, 3,4-dimethoxybenzyl carbonate, o-nitrobenzyl carbonate,p-nitrobenzyl carbonate, S-benzyl thiocarbonate, 4-ethoxy-1-napththylcarbonate, methyl dithiocarbonate, 2-iodobenzoate, 4-azidobutyrate,4-nitro-4-methylpentanoate, o-(dibromomethyl)benzoate,2-formylbenzenesulfonate, 2-(methylthiomethoxy)ethyl,4-(methylthiomethoxy)butyrate, 2-(methylthiomethoxymethyl)benzoate,2,6-dichloro-4-methylphenoxyacetate,2,6-dichloro-4-(1,1,3,3-tetramethylbutyl)phenoxyacetate, 2,4-bis(1,1-dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate,monosuccinoate, (E)-2-methyl-2-butenoate, o-(methoxyacyl)benzoate,α-naphthoate, nitrate, alkyl N,N,N′,N′-tetramethylphosphorodiamidate,alkyl N-phenylcarbamate, borate, dimethylphosphinothioyl, alkyl2,4-dinitrophenylsulfenate, sulfate, methanesulfonate (mesylate),benzylsulfonate, and tosylate (Ts).

In certain embodiments, the substituent present on a sulfur atom is asulfur protecting group (also referred to as a “thiol protectinggroup”). Sulfur protecting groups include, but are not limited to,—R^(aa), —N(R^(bb))₂, —C(═O)SR^(aa), —C(═O)R^(aa), —CO₂R^(aa),—C(═O)N(R^(bb))₂, —C(═NR^(bb))R^(aa), —C(═NR^(bb))OR^(aa),—C(═NR^(bb))N(R^(bb))₂, —S(═O)R^(aa), —SO₂R^(aa), —Si(R^(aa))₃,—P(R^(cc))₂, —P(R^(cc))₃ ⁺X⁻, —P(OR^(cc))₂, —P(OR^(cc))₃ ⁺X⁻,—P(═O)(R^(aa))₂, —P(═O)(ORC)₂, and —P(═O)(N(R^(bb))₂)₂, wherein R^(aa),R^(bb), and R^(cc) are as defined herein. Sulfur protecting groups arewell known in the art and include those described in detail inProtecting Groups in Organic Synthesis, T. W. Greene and P. G. M. Wuts,3^(rd) edition, John Wiley & Sons, 1999, incorporated herein byreference.

A “counterion” or “anionic counterion” is a negatively charged groupassociated with a positively charged group in order to maintainelectronic neutrality. An anionic counterion may be monovalent (i.e.,including one formal negative charge). An anionic counterion may also bemultivalent (i.e., including more than one formal negative charge), suchas divalent or trivalent. Exemplary counterions include halide ions(e.g., F⁻, Cl⁻, Br⁻, I⁻), NO³⁻, ClO₄ ⁻, OH⁻, H₂PO₄ ⁻, HCO₃ ⁻, HSO₄ ⁻,sulfonate ions (e.g., methansulfonate, trifluoromethanesulfonate,p-toluenesulfonate, benzenesulfonate, 10-camphor sulfonate,naphthalene-2-sulfonate, naphthalene-1-sulfonic acid-5-sulfonate,ethan-1-sulfonic acid-2-sulfonate, and the like), carboxylate ions(e.g., acetate, propanoate, benzoate, glycerate, lactate, tartrate,glycolate, gluconate, and the like), BF₄ ⁻, PF₄ ⁻, PF₆ ⁻, AsF₆ ⁻, SbF₆⁻, B[3,5-(CF₃)₂C₆H₃]₄]⁻, B(C₆F₅)₄ ⁻, BPh₄ ⁻, Al(OC(CF₃)₃)₄ ⁻, andcarborane anions (e.g., CB₁₁H₁₂ ⁻ or (HCB₁₁Me₅Br₆)⁻). Exemplarycounterions which may be multivalent include CO₃ ²⁻, HPO₄ ⁻², PO₄ ³⁻,B₄O₇ ²⁻, SO₄ ²⁻, S₂O₃ ²⁻, carboxylate anions (e.g., tartrate, citrate,fumarate, maleate, malate, malonate, gluconate, succinate, glutarate,adipate, pimelate, suberate, azelate, sebacate, salicylate, phthalates,aspartate, glutamate, and the like), and carboranes.

An “aliphatic chain” refers to a substituted or unsubstituted divalentalkyl, alkenyl, or alkynyl group. An aliphatic chain includes (1) one ormore chains of carbon atoms immediately between the two radicals of thealiphatic chain; (2) optionally one or more hydrogen atoms on thechain(s) of carbon atoms; and (3) optionally one or more substituents(“non-chain substituents,” which are not hydrogen) on the chain(s) ofcarbon atoms. A chain of carbon atoms consists of consecutivelyconnected carbon atoms (“chain atoms”) and does not include hydrogenatoms or heteroatoms. However, a non-chain substituent of an aliphaticchain may include any atoms, including hydrogen atoms, carbon atoms, andheteroatoms. For example, aliphatic chain —C^(A)H(C^(B)H₂CH₃)— includesone chain atom C^(A), one hydrogen atom on C^(A), and non-chainsubstituent −(C^(B)H₂C^(C)H₃). The term “C_(x) aliphatic chain,” whereinx is a positive integer, refers to an aliphatic chain that includes xnumber of chain atom(s) between the two radicals of the aliphatic chain.If there is more than one possible value of x, the smallest possiblevalue of x is used for the definition of the aliphatic chain. Forexample, —CH(C₂H₅)— is a C₁ aliphatic chain, and

is a C₃ aliphatic chain. When a range of values is used, the meaning ofthe range is as described herein. For example, a C₃₋₁₀ aliphatic chainrefers to an aliphatic chain where the number of chain atoms of theshortest chain of carbon atoms immediately between the two radicals ofthe aliphatic chain is 3, 4, 5, 6, 7, 8, 9, or 10. An aliphatic chainmay be saturated (e.g., —(CH₂)₄—). An aliphatic chain may also beunsaturated and include one or more C═C and/or C≡C bonds anywhere in thealiphatic chain. For instance, —CH═CH—(CH₂)₂—, —CH₂—C≡C—CH₂—, and—C≡C—CH═CH— are all examples of a unsubstituted and unsaturatedaliphatic chain. In certain embodiments, the aliphatic chain isunsubstituted (e.g., —C≡C— or —(CH₂)₄—). In certain embodiments, thealiphatic chain is substituted (e.g., —CH(C₂H₅)— and —CF₂—). Any twosubstituents on the aliphatic chain may be joined to form an optionallysubstituted carbocyclyl, optionally substituted heterocyclyl, optionallysubstituted aryl, or optionally substituted heteroaryl ring. Forinstance

are all examples of an aliphatic chain. In contrast, in certainembodiments,

are not within the scope of the aliphatic chains described herein. A“heteroaliphatic chain” is an aliphatic chain where at least one chainatom of each chain of the aliphatic chain is independently replaced witha heteroatom. In certain embodiments, an aliphatic chain describedherein is a C₁₋₄ aliphatic chain, a C₁₋₆ aliphatic chain, a C₃₋₇aliphatic chain, a C₂ aliphatic chain, a C₃ aliphatic chain, or a C₅aliphatic chain.

In certain embodiments, one, two, or three chain atoms of an aliphaticchain described herein are independently replaced with —O—, —S—, —NR¹—,—N═, or ═N—, wherein each instance of R¹ is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group.In certain embodiments, one, two, or three chain atoms of an aliphaticchain described herein are independently replaced with —O—, —S—, or—NR¹—. In certain embodiments, the molecular weight of an aliphatic orheteroaliphatic chain is not more than about 300 g/mol, not more thanabout 200 g/mol, not more than about 150 g/mol, not more than about 100g/mol, not more than about 70 g/mol, not more than about 50 g/mol, ornot more than 30 g/mol. In certain embodiments, an aliphatic orheteroaliphatic chain consists of not more than about 70 atoms, not morethan about 50 atoms, not more than about 30 atoms, not more than about20 atoms, not more than about 15 atoms, or not more than 10 atoms. Incertain embodiments, an aliphatic or heteroaliphatic chain does notinclude unsaturated bonds in the shortest chain. In certain embodiments,an aliphatic or heteroaliphatic chain consists of one or two unsaturatedbonds in the shortest chain. In certain embodiments, an aliphatic orheteroaliphatic chain includes at least one instance of ═O as anon-chain substituent on a chain atom (e.g., carbon or sulfur atom).

The term “pharmaceutically acceptable salt” refers to those salts whichare, within the scope of sound medical judgment, suitable for use incontact with the tissues of humans and lower animals without unduetoxicity, irritation, allergic response and the like, and arecommensurate with a reasonable benefit/risk ratio. Pharmaceuticallyacceptable salts are well known in the art. For example, Berge et al.,describe pharmaceutically acceptable salts in detail in J.Pharmaceutical Sciences, 1977, 66, 1-19, incorporated herein byreference. Pharmaceutically acceptable salts of the compounds of thisdisclosure include those derived from suitable inorganic and organicacids and bases. Examples of pharmaceutically acceptable, nontoxic acidaddition salts are salts of an amino group formed with inorganic acidssuch as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuricacid, and perchloric acid or with organic acids such as acetic acid,oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, ormalonic acid or by using other methods known in the art such as ionexchange. Other pharmaceutically acceptable salts include adipate,alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate,borate, butyrate, camphorate, camphorsulfonate, citrate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,formate, fumarate, glucoheptonate, glycerophosphate, gluconate,hemisulfate, heptanoate, hexanoate, hydroiodide,2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, laurylsulfate, malate, maleate, malonate, methanesulfonate,2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate,pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate,pivalate, propionate, stearate, succinate, sulfate, tartrate,thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and thelike. Salts derived from appropriate bases include alkali metal,alkaline earth metal, ammonium and N⁺(C₁₋₄ alkyl)₄ ⁻ salts.Representative alkali or alkaline earth metal salts include sodium,lithium, potassium, calcium, magnesium, and the like. Furtherpharmaceutically acceptable salts include, when appropriate, nontoxicammonium, quaternary ammonium, and amine cations formed usingcounterions such as halide, hydroxide, carboxylate, sulfate, phosphate,nitrate, lower alkyl sulfonate, and aryl sulfonate.

The term “solvate” refers to forms of the compound that are associatedwith a solvent, usually by a solvolysis reaction. This physicalassociation may include hydrogen bonding. Conventional solvents includewater, methanol, ethanol, acetic acid, DMSO, THF, diethyl ether, and thelike. The compounds described herein may be prepared, e.g., incrystalline form, and may be solvated. Suitable solvates includepharmaceutically acceptable solvates and further include bothstoichiometric solvates and non-stoichiometric solvates. In certaininstances, the solvate will be capable of isolation, for example, whenone or more solvent molecules are incorporated in the crystal lattice ofa crystalline solid. “Solvate” encompasses both solution-phase andisolatable solvates. Representative solvates include hydrates,ethanolates, and methanolates.

The term “hydrate” refers to a compound that is associated with water.Typically, the number of the water molecules contained in a hydrate of acompound is in a definite ratio to the number of the compound moleculesin the hydrate. Therefore, a hydrate of a compound may be represented,for example, by the general formula R.x H₂O, wherein R is the compound,and x is a number greater than 0. A given compound may form more thanone type of hydrate, including, e.g., monohydrates (x is 1), lowerhydrates (x is a number greater than 0 and smaller than 1, e.g.,hemihydrates (R.0.5 H₂O)), and polyhydrates (x is a number greater than1, e.g., dihydrates (R.2 H₂O) and hexahydrates (R.6 H₂O)).

The term “tautomers” or “tautomeric” refers to two or moreinterconvertible compounds resulting from at least one formal migrationof a hydrogen atom and at least one change in valency (e.g., a singlebond to a double bond, a triple bond to a single bond, or vice versa).The exact ratio of the tautomers depends on several factors, includingtemperature, solvent, and pH. Tautomerizations (i.e., the reactionproviding a tautomeric pair) may catalyzed by acid or base. Exemplarytautomerizations include keto-to-enol, amide-to-imide, lactam-to-lactim,enamine-to-imine, and enamine-to-(a different enamine) tautomerizations.

It is also to be understood that compounds that have the same molecularformula but differ in the nature or sequence of bonding of their atomsor the arrangement of their atoms in space are termed “isomers”. Isomersthat differ in the arrangement of their atoms in space are termed“stereoisomers”.

Stereoisomers that are not mirror images of one another are termed“diastereomers” and those that are non-superimposable mirror images ofeach other are termed “enantiomers”. When a compound has an asymmetriccenter, for example, it is bonded to four different groups, a pair ofenantiomers is possible. An enantiomer can be characterized by theabsolute configuration of its asymmetric center and is described by theR- and S-sequencing rules of Cahn and Prelog, or by the manner in whichthe molecule rotates the plane of polarized light and designated asdextrorotatory or levorotatory (i.e., as (+) or (−)-isomersrespectively). A chiral compound can exist as either individualenantiomer or as a mixture thereof. A mixture containing equalproportions of the enantiomers is called a “racemic mixture”.

The term “polymorphs” refers to a crystalline form of a compound (or asalt, hydrate, or solvate thereof) in a particular crystal packingarrangement. All polymorphs have the same elemental composition.Different crystalline forms usually have different X-ray diffractionpatterns, infrared spectra, melting points, density, hardness, crystalshape, optical and electrical properties, stability, and solubility.Recrystallization solvent, rate of crystallization, storage temperature,and other factors may cause one crystal form to dominate. Variouspolymorphs of a compound can be prepared by crystallization underdifferent conditions.

The term “prodrugs” refer to compounds which have cleavable groups andbecome by solvolysis or under physiological conditions the compoundsdescribed herein, which are pharmaceutically active in vivo. Suchexamples include, but are not limited to, choline ester derivatives andthe like, N-alkylmorpholine esters and the like. Other derivatives ofthe compounds described herein have activity in both their acid and acidderivative forms, but in the acid sensitive form often offer advantagesof solubility, tissue compatibility, or delayed release in the mammalianorganism (see, Bundgard, H., Design of Prodrugs, pp. 7-9, 21-24,Elsevier, Amsterdam 1985). Prodrugs include acid derivatives well knownto practitioners of the art, such as, for example, esters prepared byreaction of the parent acid with a suitable alcohol, or amides preparedby reaction of the parent acid compound with a substituted orunsubstituted amine, or acid anhydrides, or mixed anhydrides. Simplealiphatic or aromatic esters, amides, and anhydrides derived from acidicgroups pendant on the compounds described herein are particularprodrugs. In some cases it is desirable to prepare double ester typeprodrugs such as (acyloxy)alkyl esters or((alkoxycarbonyl)oxy)alkylesters. C₁-C₈ alkyl, C₂-C₈ alkenyl, C₂-C₈alkynyl, aryl, C₇-C₁₂ substituted aryl, and C₇-C₁₂ arylalkyl esters ofthe compounds described herein may be preferred.

The “molecular weight” of a monovalent moiety —R is calculated bysubtracting 1 from the molecular weight of the compound R—H. The“molecular weight” of a divalent moiety -L- is calculated by subtracting2 from the molecular weight of the compound H-L-H.

The terms “composition” and “formulation” are used interchangeably.

A “subject” to which administration is contemplated includes, but is notlimited to, humans (i.e., a male or female of any age group, e.g., apediatric subject (e.g., infant, child, adolescent) or adult subject(e.g., young adult, middle-aged adult, or senior adult)) and/or othernon-human animals, for example, mammals (e.g., primates (e.g.,cynomolgus monkeys, rhesus monkeys); commercially relevant mammals suchas cattle, pigs, horses, sheep, goats, cats, and/or dogs); and birds(e.g., commercially relevant birds such as chickens, ducks, geese,and/or turkeys). In certain embodiments, the animal is a mammal. Theanimal may be a male or female at any stage of development. The animalmay be a transgenic animal or genetically engineered animal. In certainembodiments, the subject is a non-human animal. In certain embodiments,the animal is a fish or reptile.

The term “administer,” “administering,” or “administration” refers toimplanting, absorbing, ingesting, injecting, inhaling, or otherwiseintroducing a compound or cell described herein or generated asdescribed herein, or a composition thereof, in or on a subject.

The terms “treatment,” “treat,” and “treating” refer to reversing,alleviating, delaying the onset of, or inhibiting the progress of adisease. In some embodiments, treatment may be administered after one ormore signs or symptoms of the disease have developed or have beenobserved. In other embodiments, treatment may be administered in theabsence of signs or symptoms of the disease. For example, treatment maybe administered to a susceptible subject prior to the onset of symptoms(e.g., in light of a history of symptoms and/or in light of exposure toa pathogen and/or in light of detecting that the subject has a genotypeassociated with the disease). Treatment may also be continued aftersymptoms have resolved, for example, to delay or prevent recurrence.

The terms “condition,” “disease,” and “disorder” are usedinterchangeably.

A “kinase” is a type of enzyme that transfers phosphate groups from highenergy donor molecules, such as ATP, to specific substrates, referred toas phosphorylation. Kinases are part of the larger family ofphosphotransferases. One of the largest groups of kinases are proteinkinases, which act on and modify the activity of specific proteins.Kinases are used extensively to transmit signals and control complexprocesses in cells. Various other kinases act on small molecules such aslipids, carbohydrates, amino acids, and nucleotides, either forsignaling or to prime them for metabolic pathways. Kinases are oftennamed after their substrates. More than 500 different protein kinaseshave been identified in humans. These exemplary human protein kinasesinclude, but are not limited to, AAK1, ABL, ACK, ACTR2, ACTR2B, AKT1,AKT2, AKT3, ALK, ALK1, ALK2, ALK4, ALK7, AMPKa1, AMPKa2, ANKR^(D3),ANPa, ANPb, ARAF, ARAFps, ARG, AurA, AurAps1, AurAps2, AurB, AurBps1,AurC, AXL, BAR^(K1), BAR^(K2), BIKE, BLK, BMPR1A, BMPR1Aps1, BMPR1Aps2,BMPR1B, BMPR2, BMX, BRAF, BRAFps, BRK, BRSK1, BRSK2, BTK, BUB1, BUBR1,CaMK1a, CaMK1b, CaMK1d, CaMK1g, CaMK2a, CaMK2b, CaMK2d, CaMK2g, CaMK4,CaMKK1, CaMKK2, caMLCK, CASK, CCK4, CCRK, CDC2, CDC7, CDK10, CDK11,CDK2, CDK3, CDK4, CDK4ps, CDK5, CDK5ps, CDK6, CDK7, CDK7ps, CDK8,CDK8ps, CDK9, CDKL1, CDKL2, CDKL3, CDKL4, CDKL5, CGDps, CHED, CHK1,CHK2, CHK2ps1, CHK2ps2, CK1a, CK1a2, CK1aps1, CK1aps2, CK1aps3, CK1d,CK1e, CK1g1, CK1g2, CK1g2ps, CK1g3, CK2a1, CK2a1-rs, CK2a2, CLIK1,CLIKIL, CLK1, CLK2, CLK2ps, CLK3, CLK3ps, CLK4, COT, CRIK, CR^(K7), CSK,CTK, CYGD, CYGF, DAPK1, DAPK2, DAPK3, DCAMKL1, DCAMKL2, DCAMKL3, DDR1,DDR2, DLK, DMPK1, DMPK2, DRAK1, DRAK2, DYRKIA, DYRKIB, DYR^(K2),DYR^(K3), DYR^(K4), EGFR, EphA1, EphA10, EphA2, EphA3, EphA4, EphA5,EphA6, EphA7, EphA8, EphB1, EphB2, EphB3, EphB4, EphB6, Erk1, Erk2,Erk3, Erk3ps1, Erk3ps2, Erk3ps3, Erk3ps4, Erk4, Erk5, Erk7, FAK, FER,FERps, FES, FGFR1, FGFR2, FGFR3, FGFR4, FGR, FLT1, FLT1ps, FLT3, FLT4,FMS, FRK, Fused, FYN, GAK, GCK, GCN2, GCN22, GPR^(K4), GPR^(K5),GPR^(K6), GPR^(K6)ps, GPR^(K7), GSK3A, GSK3B, Haspin, HCK, HER2/ErbB2,HER3/ErbB3, HER4/ErbB4, HH498, HIPK1, HIPK2, HIPK3, HIPK4, HPK1, HRI,HRIps, HSER, HUNK, ICK, IGF1R, IKKa, IKKb, IKKe, ILK, INSR, IRAK1,IRAK2, IRAK3, IRAK4, IRE1, IRE2, IRR, ITK, JAK1, JAK2, JAK3, JNK1, JNK2,JNK3, KDR, KHS1, KHS2, KIS, KIT, KSGCps, KSR1, KSR2, LATS1, LATS2, LCK,LIMK1, LIMK2, LIMK2ps, LKB1, LMR1, LMR2, LMR3, LOK, LRRK1, LRRK2, LTK,LYN, LZK, MAK, MAP2K1, MAP2K1ps, MAP2K2, MAP2K2ps, MAP2K3, MAP2K4,MAP2K5, MAP2K6, MAP2K7, MAP3K1, MAP3K2, MAP3K3, MAP3K4, MAP3K5, MAP3K6,MAP3K7, MAP3K8, MAPKAPK2, MAPKAPK3, MAPKAPK5, MAPKAPKps1, MARK1, MARK2,MARK3, MARK4, MARKps01, MARKps02, MARKps03, MARKps04, MARKps05,MARKps07, MARKps08, MARKps09, MARKps10, MARKps11, MARKps12, MARKps13,MARKps15, MARKps16, MARKps17, MARKps18, MARKps19, MARKps20, MARKps21,MARKps22, MARKps23, MARKps24, MARKps25, MARKps26, MARKps27, MARKps28,MARKps29, MARKps30, MAST1, MAST2, MAST3, MAST4, MASTL, MELK, MER, MET,MISR2, MLK1, MLK2, MLK3, MLK4, MLKL, MNK1, MNK1ps, MNK2, MOK, MOS,MPSK1, MPSK1ps, MRCKa, MRCKb, MRCKps, MSK1, MSK12, MSK2, MSK22, MSSK1,MST1, MST2, MST3, MST3ps, MST4, MUSK, MYO3A, MYO3B, MYT1, NDR1, NDR2,NEK1, NEK10, NEK11, NEK2, NEK2ps1, NEK2ps2, NEK2ps3, NEK3, NEK4, NEK4ps,NEK5, NEK6, NEK7, NEK8, NEK9, NIK, NIM1, NLK, NRBP1, NRBP2, NuaK1,NuaK2, Obscn, Obscn2, OSR1, p38a, p38b, p38d, p38g, p70S6K, p70S6Kb,p70S6Kps1, p70S6Kps2, PAK1, PAK2, PAK2ps, PAK3, PAK4, PAK5, PAK6, PASK,PBK, PCTAIRE1, PCTAIRE2, PCTAIRE3, PDGFRa, PDGFRb, PDK1, PEK, PFTAIRE1,PFTAIRE2, PHKg1, PHKg1ps1, PHKg1ps2, PHKglps3, PHKg2, PIK3R4, PIM1,PIM2, PIM3, PINK1, PITSLRE, PKACa, PKACb, PKACg, PKCa, PKCb, PKCd, PKCe,PKCg, PKCh, PKCi, PKCips, PKCt, PKCz, PKD1, PKD2, PKD3, PKG1, PKG2,PKN1, PKN2, PKN3, PKR, PLK1, PLK1ps1, PLK1ps2, PLK2, PLK3, PLK4, PRKX,PRKXps, PRKY, PRP4, PRP4ps, PRPK, PSKH1, PSKH1ps, PSKH2, PYK2, QIK, QSK,RAF1, RAF1ps, RET, RHOK, RIPK1, RIPK2, RIPK3, RNAseL, ROCK1, ROCK2, RON,ROR1, ROR2, ROS, RSK1, RSK12, RSK2, RSK22, RSK3, RSK32, RSK4, RSK42,RSKL1, RSKL2, RYK, RYKps, SAKps, SBK, SCYL1, SCYL2, SCYL2ps, SCYL3, SGK,SgK050ps, SgK069, SgK071, SgKO85, SgK110, SgK196, SGK2, SgK223, SgK269,SgK288, SGK3, SgK307, SgK384ps, SgK396, SgK424, SgK493, SgK494, SgK495,SgK496, SIK (e.g., SIK1, SIK2), skMLCK, SLK, Slob, smMLCK, SNRK, SPEG,SPEG2, SRC, SRM, SRPK1, SRPK2, SRPK2ps, SSTK, STK33, STK33ps, STLK3,STLK5, STLK6, STLK6ps1, STLK6-rs, SuRTK106, SYK, TAK1, TAO1, TAO2, TAO3,TBCK, TBK1, TEC, TESK1, TESK2, TGFbR1, TGFbR2, TIE1, TIE2, TLK1, TLK1ps,TLK2, TLK2ps1, TLK2ps2, TNK1, Trad, Trb1, Trb2, Trb3, Trio, TRKA, TRKB,TRKC, TSSK1, TSSK2, TSSK3, TSSK4, TSSKps1, TSSKps2, TTBK1, TTBK2, TTK,TTN, TXK, TYK2, TYK22, TYRO3, TYRO3ps, ULK1, ULK2, ULK3, ULK4, VACAMKL,VRK1, VRK2, VRK3, VRK3ps, Wee1, Wee1B, Wee1Bps, Wee1ps1, Wee1ps2, Wnk1,Wnk2, Wnk3, Wnk4, YANK1, YANK2, YANK3, YES, YESps, YSK1, ZAK, ZAP70,ZC1/HGK, ZC2/TNIK, ZC3/MINK, and ZC4/NRK.

The term “inhibition”, “inhibiting”, “inhibit,” or “inhibitor” refer tothe ability of a compound to reduce, slow, halt, or prevent activity ofa particular biological process (e.g., kinase activity) in a cellrelative to vehicle.

By “pluripotency” and pluripotent stem cells it is meant that such cellshave the ability under appropriate conditions to differentiate intocells that are derivatives of all three embryonic germ layers (endoderm,mesoderm and ectoderm). A pluripotent cell line or cell culture is oftencharacterized in that the cells can differentiate into a wide variety ofcell types in vitro and in vivo. Cells that are able to form teratomascontaining cells having characteristics of endoderm, mesoderm, andectoderm when injected into SCID mice are considered pluripotent. Inaddition, cells that possess the ability to participate in the formationof chimeras (upon injection into a blastocyst of the same species thatis transferred to a suitable foster mother of the same species) thatsurvive to term are considered pluripotent. Pluripotent cell types asused in the present invention may be provided in the form of humanembryonic stem cells, or human induced pluripotent cell (iPS cell), ormay be derived from a human embryonic stem cell line.

The term “stem cell” refers to a vertebrate cell that has the abilityboth to self-renew, and to generate differentiated progeny. The abilityto generate differentiated progeny may be described as pluripotent (seeMorrison et al. (1997) Cell 88:287-298). “Embryonic stem cells” (EScells) are pluripotent stem cells derived from the inner cell mass of ablastocyst, an early-stage preimplantation embryo. Pluripotencydistinguishes embryonic stem cells from adult stem cells found inadults; while embryonic stem cells can generate all cell types in thebody, adult stem cells are multipotent and can produce only a limitednumber of cell types.

“Induced pluripotent stem cells”, abbreviated as iPS cells, are a typeof pluripotent stem cell artificially derived from a non-pluripotentcell, typically an adult somatic cell, by inducing expression of certaingenes (e.g., injection of an expression construct). Induced pluripotentstem cells are identical in many respects to natural pluripotent stemcells, such as embryonic stem (ES) cells (e.g., in their physicalproperties). They may be the same in their expressions of certain stemcell genes and proteins, chromatin methylation patterns, doubling time,embryoid body formation, teratoma formation, viable chimera formation,and potency and differentiability. The term “induced pluripotent stemcell” encompasses pluripotent cells, that, like embryonic stem (ES)cells, can be cultured over a long period of time while maintaining theability to differentiate into all types of cells in an organism.However, unlike ES cells (which are typically derived from the innercell mass of blastocysts), iPS cells are derived from differentiatedsomatic cells, that is, cells that have a narrower, more definedpotential.

By “culturing” the cell means growing the cells in an artificial, invitro environment. By “maintaining” means continuing to grow the cellsin culture under suitable conditions until the pluripotency state of thecell is converted to a more naïve state.

A “cell culture medium” (also referred to herein as a “culture medium”or “medium”) is a medium for culturing cells containing nutrients thatmaintain cell viability and support proliferation. The cell culturemedium may contain any of the following nutrients in appropriate amountsand combinations: salt(s), buffer(s), amino acids, glucose or othersugar(s), antibiotics, serum or serum replacement, and other componentssuch as, but not limited to, peptide growth factors, cofactors, andtrace elements. Cell culture media ordinarily used for particular celltypes are known to those skilled in the art. For example, cell culturemedia of use for culturing and maintaining pluripotent cells are knownin the art.

In some embodiments, the cell culture medium is chemically definedmedium. In some embodiments, cell culture medium is serum-free medium,e.g., mTeSR1™ medium (StemCell Technologies, Vancouver, BC). In someembodiments, the culture medium comprises one or more supplements, suchas, but not limited to N2 and B27. In some embodiments, the cell culturemedium comprises a serum replacement composition. In some embodiments,the cell culture medium comprises low amount, such as less than 1% orless than 0.5%, of knock-out serum replacement medium. In someembodiments, the cell culture medium does not comprise a serumreplacement composition. In some embodiments, the cell culture mediumcomprises an activator of STAT3 pathways, for example but not limited toleukemia inhibitory factor (LIF). In some embodiments, the cell culturecomprises serum free recombinant human LIF.

In some embodiments, the cell culture medium comprises a basal medium towhich one or more supplements are added, such as: DMEM/F12, Neurobasal,N2 supplement, 10 mL B27 supplement, human LIF, glutamine, nonessentialamino acids, β-mercaptoethanol, penicillin-streptomycin, and/or BSA(Sigma). In some embodiments, the supplemented basal cell culture mediumfurther comprises fibroblast growth factor 2 (FGF2) and 1%, 0.8%, 0.6%,0.4%, 0.2%, or 0.1% KSR.

In some embodiments, the cell culture medium is free or essentially freeof components of non-human origin. In some embodiments, the cell culturemedium is free or essentially free of components isolated from humans ornon-human animals. In some embodiments, the cell culture medium usesrecombinant human proteins (e.g., recombinant human albumin).

“Cell line” refers to a population of largely or substantially identicalcells, wherein the cells have often been derived from a single ancestorcell or from a defined and/or substantially identical population ofancestor cells. For example, a cell line may consist of descendants of asingle cell. A cell line may have been or may be capable of beingmaintained in culture for an extended period (e.g., months, years, foran unlimited period of time). It will be appreciated that cells mayacquire mutations and possibly epigenetic changes over time such thatsome individual cells of a cell line may differ with respect to eachother. In some embodiments, at least 80%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the cells of a cellline or cell culture are at least 95%, 96%, 97%, 98%, or 99% geneticallyidentical. In some embodiments, at least 80%, 85%, 86%, 87%, 88%, 89%,90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% of the cells of acell line or cell culture express the same set of cell surface markers.The set of markers could be markers indicative of ground state (naïve)pluripotency or cell-type specific markers.

A “clone” refers to a cell derived from a single cell without change. Itwill be understood that if cells of a clone are subjected to differentculture conditions or if some of the cells are subjected to geneticmodification, the resulting cells may be considered distinct clones.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a reporter system for naïve human pluripotency based onendogenous OCT4 distal enhancer activity. In FIG. 1A, the proximalenhancer (PE) targeting strategy in human ESCs containing a 2A-GFPsequence in frame with the 3′ UTR of OCT4 is shown. FIG. 1B is aSouthern blot analysis confirming disruption of the PE in OCT4-GFP ESCs.NdeI-digested genomic DNA was hybridized with 5′ and 3′ external probes.Expected fragment size: WT (wild type)=5.6 kb, T (targeted)=6.4 kb. FIG.1C shows the images of OCT4-GFP human ESCs before (left) and afterTALEN-mediated deletion of the PE (right). FIG. 1D shows the singlemolecule RNA FISH analysis for OCT4 and GFP transcripts in OCT4-GFPhuman ESCs before and after TALEN-mediated disruption of the PE. FIG. 1Eshows the flow cytometric analysis of the proportion of OCT4-ΔPE-GFP+cells obtained after DOX induction of lentiviral KLF2, NANOG orKLF2+NANOG. Following primary infection WIBR3 human ESCs containing theOCT4-ΔPE-GFP reporter allele were trypsinized and treated with hESM,2i/L, or 2i/L/DOX for one week. FIG. 1F are phase and fluorescenceimages and flow cytometric analysis of the proportion of OCT4-ΔPE-GFP+cells of a clonal line of 38 hESCs derived in 2i/L/DOX (left). Phase andfluorescence images and flow cytometric analysis after replating in theabsence of DOX for one week (right). FIG. 1G shows the quantitative geneexpression analysis for FUW-KLF2, FUW-NANOG, endogenous OCT4, andendogenous KLF4 in WIBR3 hESCs cultured in hESM and clonal OCT4-ΔPE-GFP+derivatives generated in 2i/L/DOX. FIG. 1H are phase and fluorescenceimages of primitive neural stem cells (pNSCs) derived by treating WIBR3hESCs containing the OCT4-ΔPE-GFP allele with 2i/L for three passages.FIG. 1I shows immunofluorescence staining for NANOG and NESTIN in aclonal line of OCT4-ΔPE-GFP-positive cells derived in 2i/L/DOX, and aclonal line of OCT4-ΔPE-GFP-negative pNSCs derived in 2i/L. FIG. 1J is amodel representing the distinct phenotypic responses of hESCs totreatment with 2i/L and 2i/L/DOX. OCT4-ΔPE-GFP+ cells generated in2i/L/DOX do not maintain reporter activity upon transgene withdrawal.OCT4-ΔPE-GFP+ cells can revert back to the conventional ‘primed’ hESCstate by re-exposure to serum and FGF.

FIG. 2 shows the identification of small molecules that maintainOCT4-ΔPE-GFP activity after transgene withdrawal. FIG. 2A shows thestrategy for screening a kinase inhibitor library to identify compoundsthat maintain OCT4-ΔPE-GFP reporter activity upon withdrawal ofDOX-dependent KLF2 and NANOG expression. FIG. 2B is the raw dataobtained from high-throughput flow cytometric analysis of the proportionof OCT4-ΔPE-GFP+ cells in 96 wells supplemented with a kinase inhibitorlibrary (n=2). FIG. 2C shows the hit compounds from a maintenance screenusing a clonal line of WIBR3 OCT4-ΔPE-GFP+ ESCs established in 2i/L/DOX.FIG. 2D are the phase images (top) and flow cytometric analysis of theproportion of OCT4-ΔPE-GFP+ cells (bottom) in a clonal line ofOCT4-ΔPE-GFP+ cells derived in 2i/L/DOX and maintained for 10 passageswithout DOX in the presence of each candidate compound. FIG. 2E is thequantitative gene expression analysis for FUW-KLF2, FUW-NANOG,endogenous OCT4, and EGFP in a clonal line of OCT4-ΔPE-GFP+ cellsmaintained in 2i/L/DOX or for five passages without DOX in the presenceof each candidate compound. FIG. 2F is the chemical structure of theBRAF inhibitor, SB590885. FIG. 2G shows phase images of a clonal line ofWIBR3 human ESCs established in 2i/L/DOX and maintained for 8 passageswithout DOX in 2i/L/SB590885 (1 μM). FIG. 2H is the quantitative geneexpression analysis for FUW-KLF2, FUW-NANOG, and endogenous OCT4 in twoclonal lines of WIBR3 human ESCs maintained for 8 passages without DOXin 2i/L/SB590885 (1 μM).

FIG. 3 shows the optimization of medium for maintaining viableOCT4-ΔPE-GFP+ cells.

FIG. 3A shows the flow cytometric analyses of the proportion ofOCT4-ΔPE-GFP+ cells in a 96 well one week after culture in 2i/L/DOX,2i/L alone or 2i/L/SB590885 (1 μM). Top panel shows quantification ofOCT4-ΔPE-GFP+ cells without including live/dead discrimination. Bottompanel shows quantification of OCT4-ΔPE-GFP+ cells after gating out DAPI+cells. FIG. 3B shows the strategy for screening a kinase inhibitorlibrary to identify compounds that improve the fraction of viable(DAPI−) OCT4-ΔPE-GFP+ cells maintained without DOX for 2 passages in2i/L/SB590995 (1 μM). FIG. 3C is the raw data obtained fromhigh-throughput flow cytometric analysis of the proportion ofDAPI-/OCT4-ΔPE-GFP+ cells in 96 wells supplemented with one plate of akinase inhibitor library (n=2). Hit compound WH-4-023 is indicated withchemical structure. FIGS. 3D-3E show high-throughput flow cytometricquantification of the proportion of DAPI-/OCT4-ΔPE-GFP+ cells in 96wells cultured for one passage (FIG. 3D) or two passages (FIG. 3E) in 64different concentrations of PD0325901, CHIR99021, and SB590885. All thelegends in FIG. 3E also apply in FIG. 3D. Asterisk denotes rank of thestandard concentration of the three inhibitors used in the precedingexperiments (1 μM PD0325901, 0.3 μM CHIR99021 and 0.5 μM SB590885). FIG.3F are the phase and fluorescence images (top) and flow cytometricanalysis of the proportion of OCT4-ΔPE-GFP+ cells (bottom) in a clonalline of OCT4-ΔPE-GFP+ cells derived in 2i/L/DOX and maintained for 2passages without DOX in 2i/L/SB590885^(opt)/Y-27632 or2i/L/SB590885^(opt)/Y-27632/WH-4-023. Opt=optimized concentrations ofPD0325901, CHIR99021, and SB590885 (see FIG. 4E). FIG. 3G are the phaseand fluorescence images of a clonal line of WIBR3 OCT4-ΔPE-GFP+ cells(left) and a clonal line of wild-type WIBR3 human ESCs generated in2i/L/DOX (right) and maintained for 3 passages inPD0325901/IM12/SB590885/Y-27632/WH-4-023 (5i) and hLIF. FIG. 3H showsthe teratoma generated from wild-type WIBR3 human ESCs maintained inPD0325901/IM12/SB590885/Y-27632/WH-4-023 (5i) and hLIF after transgenewithdrawal. Representative tissues of the three germ layers areindicated.

FIG. 4 shows the direct conversion of conventional human ESCs to naïvepluripotency in 5i/L. FIG. 4A shows the strategy for assessing directconversion of primed human ESCs into OCT4-ΔPE-GFP+ cells under optimizedchemical conditions. FIG. 4B shows phase and fluorescence images ofemerging naïve colony and expanded cells from WIBR3 OCT4-ΔPE-GFP humanESCs treated with 5i/L for 10 days. FIG. 4C are the phase andfluorescence images and flow cytometric analyses of the proportion ofGFP+ cells during conversion experiments in 5i/L supplemented with FGFand/or Activin A (FA). FIG. 4D are the phase images of wild-type naïveWIBR2 human ES cells converted in 5i/L supplemented with FGF and/orActivin A (FA).

FIG. 4E shows the phase image of a primary human ESC line derived in5i/L/FA conditions from an explanted human blastocyst. Cell line iscalled Whitehead Institute Naïve Human ESC (WIN)-1. FIG. 4F is the flowcytometric analysis of the proportion of OCT4-ΔPE-GFP+ cells threepassages after withdrawal of individual inhibitors and growth factors.FIG. 4G is the quantitative gene expression analysis for OCT4, NANOG,KLF4, KLF2, SOX2, and REX1 three passages after withdrawal of individualinhibitors and growth factors.

FIG. 5 shows the evaluation of alternative culture conditions for naïvehuman pluripotency. FIG. 5A is a table comparing the components of fourrecent protocols for capturing naïve-like human ESCs with 5i/L/A medium.FIG. 5B show the phase and fluorescence images and flow cytometricanalyses showing the response of OCT4-ΔPE-GFP-negative primed cells torecently reported protocols for naïve human pluripotency (see FIG. 5A)and 5i/L/A. FIG. 5C shows the quantification of the proportion ofGFP-positive cells in WIBR3 OCT4-GFP and OCT4-ΔPE-GFP human ESCs uponremoval of DOX-inducible KLF2 and NANOG expression in primed medium (PM)and four alternative conditions for naïve human pluripotency. FIG. 5Dshows the flow cytometric analysis of the proportion of OCT4-ΔPE-GFP+cells in 5i/L/A and the JNK inhibitor SP600125 (6i/L/A) in serum-freeN2B27 basal medium vs. 20% KSR basal medium. FIG. 5E is the quantitativegene expression analysis for OCT4, SOX2, KLF2 and NANOG in human ESCscultured in 6i/L/A and supplemented with 1-10% FBS or KSR. FIG. 5F arethe phase and fluorescence images of induction of OCT4-ΔPE-GFP activityfrom the primed state in 6i/L/A, and 6i/L/A supplemented with 0.5-1%KSR.

FIG. 6 shows the transcriptional profiling of naïve human ESCs in5i/L/A. FIG. 6A shows the cross-species hierarchical clustering of naïveand primed pluripotent cells from mice and humans, as performedpreviously by Gafni et al. (2013). Affymetrix expression data werenormalized using RNA spike-in. Two groups of human ESC samples areincluded: WIBR2, WIBR3 and WIN1 human ESCs derived in the optimizednaïve medium (5i/L/A or 6i/L/A, as indicated), and parental WIBR2 andWIBR3 human ESCs in primed human ESC medium. Correlation matrix of geneexpression was clustered using Pearson correlation coefficients (PCCs).The average linkage hierarchical clustering of the Pearson correlationis shown in the heatmap. mEpiSCs, mouse EpiSCs; mESC, mouse ESC; miPSC,mouse iPSC.

FIG. 6B shows the gene ontology (GO) analysis showing up- anddown-regulated gene categories with most significant p values betweenthe naïve human conditions and primed human ESCs. FIG. 6C is a volcanoplot showing fold change (x axis) between the naïve human ESC samplesand primed human ESCs on all genes (left); volcano plot showingpreviously published fold change (x axis) between the naïve human ESCsamples of Gafni et al. (2013) and primed human ESCs on all genes(right). The open circles are those that are log 2 fold change >1 and<-1 & meet a p<0.05. FIG. 6D shows the fold changes in expression ofnaïve pluripotency-associated transcripts in the naïve human ESC samplesvs. primed hESCs (blue or black), and the naïve human samples publishedby Gafni et al. (2013) vs. primed human ESCs (red or grey). FIG. 6Eshows, for comparison with (FIG. 6D), fold changes in expression ofnaïve pluripotency-associated transcripts in naïve mouse ESCs vs. primedmouse EpiSCs were curated from a previously published study (Najm etal., 2011). FIG. 6F shows the quantitative gene expression analysis forNANOG and STELLA in human ESCs cultured in parallel in primed medium,the medium of Gafni et al. (2013) and 5i/L/FA. FIG. 6G shows singlemolecule (sm) RNA FISH analysis using OCT4 and NANOG probes in WIBR2human ESCs cultured in primed medium, the medium of Gafni et al. (2013)or 5i/L/A. FIG. 6H shows single molecule (sm) RNA FISH analysis usingOCT4, NANOG, KLF4 and REX1 probes in human ESCs cultured in primedmedium, the medium of Gafni et al. (2013) or 5i/L/A.

FIG. 7 shows the chromatin landscape of naïve human pluripotency. FIG.7A-7E show the ChIP-Seq tracks for H3K4me3 and H3K27me3 at four classesof genes: (FIG. 7A) developmental genes that are bivalent in the primedstate and exhibit loss of H3K27me3 in the naïve state; (FIG. 7B) naïvepluripotency genes that are bivalent in the primed state and exhibitloss of H3K27me3 in the naïve state; (FIG. 7C) naïve pluripotency genesthat acquire H3K4me3 in the naïve state; (FIG. 7D) master transcriptionfactors that have a signal for H3K4me3, but not H3K27me3, in both naïveand primed states; (FIG. 7E) ChIP-Seq tracks for H3K4me3 and H3K27me3 atthe DUSP6 and SOX11 loci in WIBR2 human ESCs cultured under primed (redor grey) or naïve 6i/L/A (blue or black) conditions. FIG. 7F shows theChIP-Seq analysis for H3K4me3 and H3K27me3 at Polycomb target genes inWIBR2 human ESCs cultured in primed medium (left) or naïve 6i/L/A medium(right). FIG. 7G shows average H3K4me3 and H3K27me3 signal at Polycombtarget genes in WIBR2 human ESCs cultured in primed medium (red or grey)or naïve 6i/L/A medium (blue or black).

FIG. 8 shows a reporter system for naïve human pluripotency based onendogenous OCT4 distal enhancer activity (associated with FIG. 1). FIG.8A: southern blot analysis confirming deletion of the PE in OCT4-GFPESCs and the removal of floxed PGK-puro cassette. NdeI-digested genomicDNA was hybridized with 5′ and 3′ external probes. Expected fragmentsize: WT (wild type)=5.6 kb, T (targeted)=6.4 kb, PEKO (targeted alleleafter PGK-puro removal)=4.6 kb. FIG. 8B: phase and GFP images ofOCT4-ΔPE-GFP+ cells obtained after DOX induction of lentiviralKLF2+NANOG. Following primary infection, WIBR3 human ESCs containing theOCT4-ΔPE-GFP reporter allele were trypsinized and treated with hESM,2i/L or 2i/L/DOX for one week. FIG. 8C: immunofluorescence staining forOCT4 in a clonal line of OCT4-ΔPE-GFP-positive cells derived in2i/L/DOX, and a clonal line of OCT4-ΔPE-GFP-negative pNSCs derived in2i/L. FIG. 8D: quantitative gene expression analysis for EGFP, SOX2,PRMD14 and PAX6 in clonal OCT4-ΔPE-GFP+ human ESC lines generated in2i/L/DOX, secondary primed cells generated by withdrawal of DOX andexpansion in conventional hESM, and clonal lines ofOCT4-ΔPE-GFP-negative pNSCs derived in 2i/L.

FIG. 9 shows the identification of small molecules that maintainOCT4-ΔPE-GFP activity after transgene withdrawal (associated with FIG.2). FIG. 9A: raw data obtained from high-throughput flow cytometricanalysis of the proportion of OCT4-ΔPE-GFP+ cells in 96 wellssupplemented with a kinase inhibitor library (n=2). FIG. 9B:quantitative gene expression analysis for STELLA, KLF4, PRDM14 and SOX2in a clonal line of OCT4-ΔPE-GFP+ cells maintained in 2i/L/DOX or forfive passages without DOX in the presence of each candidate compound.FIG. 9C: flow cytometric analyses of the proportion of viable(DAPI-negative) and OCT4-ΔPE-GFP+ cells in 2i/L/DOX or two passagesafter DOX withdrawal in 2i/L/SB590885 (1 μM).

FIG. 10 shows the optimization of medium for maintaining viableOCT4-ΔPE-GFP+ cells (associated with FIG. 3). FIG. 10A: raw dataobtained from high-throughput flow cytometric analysis of the proportionof DAPI-/OCT4-ΔPE-GFP+ cells in 96 wells supplemented with three platesof a kinase inhibitor library in the presence of the primary hitcompound SB590885 (n=2). FIG. 10B: hit compounds from a viability screenusing a clonal line of WIBR3 OCT4-ΔPE-GFP+ ESCs established in 2i/L/DOX.FIG. 10C: phase images and flow cytometric analyses of OCT4-ΔPE-GFP+cells maintained in 2i/L/SB590885 (0.5 μM)±ROCK inhibitor Y-27632 (10μM) for two passages. FIG. 10D: phase and GFP images of OCT4-ΔPE-GFP+cells maintained for four passages in 2i/L/DOX, 2i/L/SB590885 (0.5μM)+ROCK inhibitor Y-27632 (10 μM) or the same medium in which CHIR99021was replaced with an alternative GSK3 inhibitor, IM-12 (1.0 μM).

FIG. 11 shows the direct conversion of conventional human ESCs to naïvepluripotency in 5i/L [associated with FIG. 4]. FIG. 11A: karyotypeanalysis of naïve human ESCs WIBR2 (P8 in 5i/L/A). Cytogenetic analysiswas performed on 20 metaphase cells. FIG. 11B: karyotype analysis ofnaïve human ESCs WIN1 (P7 in 5i/L/FA). Cytogenetic analysis wasperformed on 20 metaphase cells. FIG. 11C: phase images of OCT4-ΔPE-GFP+cells three passages after withdrawal of individual inhibitors andgrowth factors.

FIG. 12 shows the evaluation of alternative culture conditions for naïvehuman pluripotency [associated with FIG. 5]. FIG. 12A: phase andfluorescence images (Top) and flow cytometric analysis of the proportionof OCT4-ΔPE-GFP+ cells (Bottom) in OCT4-ΔPE-GFP+ cells derived in5i/L/FA and maintained for three passages in the presence of additionalmedia additives, as indicated. FIG. 12B: quantitative gene expressionanalysis for KLF2 and KLF4 in OCT4-ΔPE-GFP-positive naïve human ESCscultured in 5i/L/A and supplemented with various components of themedium of Gafni et al. (2013). FIG. 12C: phase and fluorescence images(Top) and flow cytometric analysis of the proportion of OCT4-ΔPE-GFP+cells (Bottom) in OCT4-ΔPE-GFP+ cells derived in 5i/L/FA and maintainedfor three passages in 20% KSR basal medium supplemented with the mediaadditives described in (FIG. 12A).

FIG. 13 shows the transcriptional profiling of naïve human ESCs in5i/L/A and 6i/L/A [associated with FIG. 6]. FIG. 13A: quantitative geneexpression analysis for KLF4 and REX1 in human ESCs cultured in parallelin primed medium, 5i/L/FA and the medium of Gafni et al. (2013). FIG.13B: expression profile of transcripts upregulated in 6i/L/A duringhuman embryonic development. For each gene, the normalized expressionvalues in human ESCs cultured in 6i/L/A vs. primed human ESCs areindicated (Left). An unpaired two-tailed t test was performed toestablish the degree of significance. Expression of the correspondingtranscript is shown at nine stages of human pre-implantationdevelopment, as detected by single cell RNA-Seq profiling (Yan et al.,2013) (Right). This comparison indicates that naïve-associatedtranscripts upregulated in 6i/L/A are enriched at the morula/epiblaststage of human development when compared to human ESCs at passage 0 orpassage 10. FIG. 13C: variability in NANOG expression compared betweensingle human ESCs cultured in primed medium, 5i/L/A or the medium ofGafni et al. (2013).

FIG. 14 shows the developmental potential of naïve human ESCs in 5i/L/A.FIG. 14A: teratomas generated from WIBR3 OCT4-ΔPE-GFP-positive humanESCs derived and maintained in 5i/L±FA and WIBR3 AAVS1-tdTomato humanESCs in 5i/L/FA. Representative tissues of the three germ layers areindicated. FIG. 14B: immunofluorescence staining for AFP and HNF4afollowing 20d of hepatic differentiation in naïve WIBR2 human ESCsderived and maintained in 5i/L/A. FIG. 14C: table summarizing injectionsof human ESCs maintained in 5i/L/FA (top) or the medium of Gafni et al.(2013) (bottom) in mouse embryos. C1-AAVS1-GFP human ESCs in the mediumof Gafni et al. (2013) were cultured on MEFs, gelatin/vitronectin ormatrigel prior to injection. (*) E10.5 embryos injected with human ESCscultured in 5i/L/FA and the medium of Gafni et al. (2013) were mixedduring collection, but none were identified as positive. (**) 30injected embryos were lost during transfer.

FIG. 15 shows that the proliferation of naïve human embryonic stem cellsis enhanced by reduction or removal of GSK3 inhibition. FIG. 15A:transgene-dependent naïve human embryonic stem (ES) cell line used totitrate the concentrations of inhibitors in 5i/L/A (see Theunissen etal., Cell Stem Cell, 2014). This subclone of WIBR3 is dependent onDoxycycline (DOX) to maintain expression of two lentiviral transgenes,KLF2 and NANOG. Left, phase image shows colony morphology in 2i/L/DOX.Middle, flow cytometric quantification shows the proportion of cellspositive for the OCT4-ΔPE-GFP reporter in 2i/L/DOX. Right, this cellline has a normal (46,XX) karyotype. FIG. 15B: titration assay to attainoptimal concentrations of small molecule inhibitors for maintenance ofnaïve human ES cells. Top, phase images showing colony morphology aftersequential passaging by single cell dissociation at low density (1:10)in four different conditions: 2i/L, 5i/L/A, t5i/L/A (0.2 μM GSK3inhibitor IM12) and 4i/L/A (removal of GSK3 inhibitor IM12). Bottom,flow cytometric quantification shows the proportion of cells positivefor the OCT4-ΔPE-GFP reporter in each condition. FIG. 15C: quantitativeRT-PCR (qRT-PCR) analysis confirming the downregulation of exogenousKLF2 and NANOG transgenes and primed marker VIMENTIN and upregulation ofnaïve markers KLF4, REX1 and STELLA in t5i/L/A and 4i/L/A. Error barsindicate one standard deviation.

FIG. 16 shows the quantification of the proliferation of naïve humancells in presence of different Wnt signal modulators. Quantification ofcell number in naïve culture conditions supplemented with distinct Wntsignal modulators after withdrawal of DOX from transgene-dependent naïvehuman ES cells (see FIG. 15A). 1×105 cells were seeded per individualwell of a 6 well plate and cells were dissociated and re-seeded at 1:5density for three successive passages. Cell numbers at successivepassages were recorded in six conditions (n=2): 2i/L/DOX (control),2i/L, 5i/L/A, 4i/L/A (removal of GSK3 inhibitor IM12), 4i/L/A+CHIR99021(1 μM), and 4i/L/A+IWR1 (2.5 μM). At passages 2 and 3 the proliferationwas significantly elevated in 4i/L/A compared to 5i/L/A. Replacing IM 12with alternative GSK3 inhibitor CHIR99021 or addition of the Wntinhibitor IWR1 did not further stimulate the proliferation of naïvehuman cells.

FIG. 17 shows the induction of naïve human pluripotency in 5i/L/A or4i/L/A is associated with X chromosome reactivation. FIG. 17A: areporter system for X chromosome status of human ES cells was engineeredby TALEN-mediated targeting of both alleles of the X-linked MECP2 genewith fluorescent reporters. FIG. 17B: starting from a single color(GFP-positive) primed line, conversion to the naïve state in 5i/L/A or4i/L/A (-IM12) results in activation of the tdTomato-labelled allelewhile GFP activity is maintained. FIG. 17C: starting from a single color(tdTomato-positive) primed line, conversion to the naïve state in 5i/L/Aor 4i/L/A (-IM12) results in activation of the GFP-labelled allele whiletdTomato activity is maintained. These results indicate that inductionof naïve human pluripotency is accompanied by a switch towards biallelicexpression of X-linked genes, which is confirmed by RNA FISH data (notshown).

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure, in one aspect, is based on the identification ofcompounds (e.g., compounds of any one of Formulae (A) to (L), andpharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof) that support self-renewal of naïve human ESCs. Inparticular, iterative chemical screening identified kinase inhibitorsthat induce and maintain OCT4 distal enhancer activity, a molecularsignature of ground state pluripotency, when applied directly toconventional human ESCs. These inhibitors generate a homogeneouspopulation of human pluripotent stem cells in which transcriptionfactors associated with the ground state of pluripotency are highlyupregulated. Comparison with previously reported naïve human ESCsindicates that the procedures defined herein capture a novel pluripotentstate in humans that closely resembles mouse ESCs. Accordingly, aspectsof the disclosure provide methods for converting the pluripotency stateof a vertebrate (e.g., human) cell to a more naïve state. Also providedare naïve pluripotent vertebrate cells (e.g., human) and cell linesproduced by the methods described herein. Some aspects of the inventionalso involve kits for converting the pluripotency state of a vertebrate(e.g., human) cell to a more naïve state.

Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs)have attracted much attention because of their potential to mature intovirtually any cell type in the body. However, mouse ESCs and iPSCs havedifferent growth factor requirements and provide a more reliable vehiclefor directed differentiation as compared to human ESCs and iPSCs. It wasthought for many years that these differences reflected variationbetween species. In 2007, however, two groups reported that novel stemcell lines derived from the post-implantation epiblast of mouse embryos,called EpiSCs, have properties similar to human ESCs (Brons et al.,2007; Tesar et al., 2007). These include a flat morphology, dependenceon bFGF and activin signaling, and use of the OCT4 proximal enhancerelement. The inner cell mass (ICM)-like state of mouse ESCs wasdescribed as “naïve,” whereas EpiSCs and human ESCs were designated as“primed” (Nichols and Smith, 2009); the implication is that the primedstate is prone to differentiate, whereas the naïve condition correspondsto the more immature “ground state” of pluripotency.

The present disclosure relates to the discovery that conventional humanESCs can be converted into a more immature/less restricted state (i.e.,a more “naïve” pluripotent state) that extensively shares definingfeatures with pluripotent mouse ESCs. Described herein are optimizedconditions that enable the interconversion between conventional andnaïve human ESCs in the absence of reprogramming factors, as well as thedirect isolation of genetically unmodified naïve ESCs from humanblastocysts. The methods described herein capture a distinct and novelstate of human pluripotency that shares defining features with mouseESCs.

Accordingly, some aspects of the disclosure provide a method forchanging the pluripotency state of a vertebrate cell to a more naïvestate. The method comprises (a) culturing a pluripotent vertebrate cellin the presence of a serine/threonine-protein kinase B-Raf (BRAF)inhibitor, an epidermal growth factor receptor (EGFR) inhibitor, avascular endothelial growth factor 1 (VEGFR1) inhibitor, or a fibroblastgrowth factor receptor 1 (FGFR1) inhibitor; and (b) maintaining the cellin culture under conditions suitable and a time sufficient to changingthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the original vertebrate cell of step (a).

The kinase inhibitor(s) used herein may be a small molecule,antibodies/antibody fragments (at least for those kinases that have anextracellular domain), short interfering RNA, and/or aptamers. A “smallmolecule,” (M) as used herein, refers to an alkyl, alkenyl, alkynyl,aryl, heteroaryl, carbocyclic, or heterocyclic moiety, as definedherein, comprising carbon and hydrogen, and optionally comprising one ormore heteroatoms as a part of the molecule (in the case of heteroaryland heterocyclic groups) and/or attached to the molecule selected fromoxygen, nitrogen, sulfur, phosphorus, boron, silicon, and selenium. Incertain embodiments, the specificity of the inhibitors is given by theIC50 value. The IC50 value is defined as the concentration of inhibitorrequired to inhibit 50% of the kinase activity. In certain embodiments,the compounds of Formula (I) or (II) may exhibit IC50 values<100 μM. Incertain other embodiments, the compounds exhibit IC50 values<50 μM. Incertain other embodiments, the compounds exhibit IC50 values<40 μM. Incertain other embodiments, the compounds exhibit IC50 values<30 μM. Incertain other embodiments, the compounds exhibit IC50 values<20 μM. Incertain other embodiments, the compounds exhibit IC50 values<10 μM. Incertain other embodiments, the compounds exhibit IC50 values<7.5 μM. Incertain embodiments, the compounds exhibit IC50 values<5 μM. In certainother embodiments, the compounds exhibit IC50 values<2.5 μM. In certainembodiments, the compounds exhibit IC50 values<1 μM. In certainembodiments, the compounds exhibit IC50 values<0.75 μM. In certainembodiments, the compounds exhibit IC50 values<0.5 μM. In certainembodiments, the compounds exhibit IC50 values<0.25 μM. In certainembodiments, the compounds exhibit IC50 values<0.1 μM. In certain otherembodiments, the compounds exhibit IC50 values<75 nM. In certain otherembodiments, the compounds exhibit IC50 values<50 nM. In certain otherembodiments, the compounds exhibit IC50 values<25 nM. In certain otherembodiments, the compounds exhibit IC50 values<10 nM. In otherembodiments, the compounds exhibit IC50 values<7.5 nM. In otherembodiments, the compounds exhibit IC50 values<5 nM.

In certain embodiments, the BRAF inhibitor is a compound of Formula (B)(e.g., AZ-628 or BAY-439006) or Formula (C) (e.g., GDC-0879 orSB590885). In certain embodiments, the BRAF inhibitor is sorafenib;PLX4720; PLX-3603; GSK2118436; N-(3-(5-(4-chlorophenyl)-1H-pyrrolo[2;3-b]pyridine-3-carbonyl)-2;4-difluorophenyl)propane-1-sulfonamide;vemurafenib (also known as Zelobraf® and PLX-4032); GSK 2118436; RAF265(Novartis); XL281; ARQ736; a compound described in international PCTapplication publication, WO 2007/002325, WO 2007/002433, WO 2009/111278,WO 2009/111279, WO 2009/111277, WO 2009/111280, or WO 2011/025927; or acompound described in U.S. Pat. No. 7,491,829 or 7,482,367. In certainembodiments, the VEGFR1 inhibitor is a compound of Formula (A) (e.g.,AMG706), Formula (B) (e.g., BAY 73-4506), or Formula (G) (e.g.,BIBF-1120 or SU11248). In certain embodiments, the VEGFR1 inhibitor isSU5416 or a compound described in U.S. patent application publication,US 2006/0030000. In certain embodiments, the FGFR1 inhibitor is acompound of Formula (F) (e.g., PD173074). In certain embodiments, theFGFR1 inhibitor is cediranib; brivanib; TSU-68; BIBF1120; dovitinib;Ki23057; MK-2461; E7080; SU5402; BGJ398; E-3810; AZD4547; PLX052; or acompound described in U.S. Pat. No. 8,709,718. In certain embodiments,the MEK inhibitor is a compound of Formula (K) (e.g., PD0325901). Incertain embodiments, the MEK inhibitor is a compound described ininternational PCT application publication, WO 2010/138377, WO2009/153554, WO 2009/093009, WO 2009/013462, WO 2009/093013, WO2008/020206, WO 2008/078086, WO 2008/120004, WO 2008/125820, WO2009/093008, WO 2009/074827, WO 2009/093009, WO 2010/108652, WO2010/105110, WO 2010/105082, WO 2009/129246, WO 2009/018238, WO2009/018233, WO 2008/089459, WO 2008/124085, WO 2008/076415, WO2008/021389, WO 2010/051935, WO 2010/051933, WO 2009/129938, WO2009/021887, WO 2008/101840, WO 2008/055236, WO 2010/003025, WO2010/003022, WO 2007/096259, WO 2008/067481, WO 2008/024724, WO2008/024725, or WO 2010/0145197; or a compound described in U.S. patentapplication publication, US 2008/0255133, US 2008/0058340, US2009/0275606, or US 2009/0246198. In certain embodiments, the GSK3inhibitor is a compound of Formula (C) (e.g., CHIR99021) or Formula (J)(e.g., IM12). In certain embodiments, the GSK3 inhibitor is CHIR98014;CHIR98023; BlO-acetoxime; BIO; LiCl; SB 216763; SB 415286; AR-A014418;1-azakenpaullone; bis-7-indolylmaleimide; kenpaullone; CT 99021; CT20026; SB216763; SB 415286; TDZD-8; TIBPO(2-thio(3-iodobenzyl)-5-(1-pyridyl)-[1,3,4]-oxadiazole); a compounddescribed in U.S. patent application publication, US 2013/0059385, US2001/0034051, US 2002/0156087, US 2004/0092535, US 2004/0209878, US2004/0138273, US 2004/0077707, US 2005/0054663, US or 2006/0089369; acompound described in U.S. Pat. No. 6,057,117; U.S. Pat. No. 6,608,063;U.S. Pat. No. 6,417,185; U.S. Pat. No. 6,489,344; or U.S. Pat. No.6,153,618; or a compound described in international PCT applicationpublication, WO/2003/049739; WO/2002/085909; WO/2003/011287,WO/2005/039485, or WO/2006/091737. In certain embodiments, the ROCKinhibitor is a compound of Formula (L) (e.g., Y-27632). In certainembodiments, the ROCK inhibitor is fasudil (HA-1077); thiazovivin;AMA0076; AR-12286; AMA0076; AR-12286; AR-13324; ATS907; DE-104;INS-115644; INS-117548; K-115; PG324; Y-39983; RKI-983; SNJ-1656; acompound described in international PCT application publication, WO2014/068035, WO 2013/030216, WO 2013/030367, WO 2013/030366, WO2013/030365, WO 2011/107608, WO 2012/146724, WO 2006/137368, or WO2005/035506; or a compound described in U.S. patent applicationpublication, US 2013/196437. In certain embodiments, the Src inhibitoris a compound of Formula (H) (e.g., WH-4-023). In certain embodiments;the Src inhibitor is SKI606 (bosutinib); dasatinib (SPYRCEL);saracatenib (AZD-0530); PP1; PP2; PD173955; AGL1872; PD162531; radicicolR2146; geldanamycin; or a compound described in U.S. patent applicationpublication, US 2006/258686, US 2009/0227608, US 2010/0249152, or US2013/0040972. ESCs are pluripotent cells. ESCs have been derived fromvertebrate animals such as mice, primates (including humans), and someother species. ESCs are often derived from cells obtained from the innercell mass (ICM) of a vertebrate blastocyst but can also be derived fromsingle blastomeres (e.g., removed from a morula). Pluripotent cells canalso be obtained using parthenogenesis, e.g., from germ cells, e.g.,oocytes. Other pluripotent cells include embryonic carcinoma (EC) andembryonic germ (EG) cells. See, e.g., Yu J, Thomson J A, Genes Dev.Pluripotent stem cell lines. 22(15):1987-97, 2008.

Standard techniques for preparing deriving human ES cells typicallyinvolve the use of a MEF or human cell feeder layer and serum or, ifcultured in serum-free medium, compounds such as bFGF. For example, theICM of a human blastocyst is removed by immunosurgery, dissociated inCa²⁺-Mg²⁺-free medium, and plated over mouse embryonic fibroblasts orhuman feeder cells (Thomson et al., Science 282, 1145 (1998). The mousecells are irradiated to suppress their proliferation. See, e.g., B. E.Reubinoff et al., Nature Biotechnol. 18, 399, 2000; Mitalipova M &Palmarini G. Isolation and characterization of human embryonic stemcells. Methods Mol. Biol. 331:55-76, 2006; Ilic D, et al., Derivation ofhESC from intact blastocysts. Curr Protoc Stem Cell Biol., Chapter1:Unit 1A.2, 2007; Ludwig T, A Thomson J., Defined, feeder-independentmedium for human embryonic stem cell culture. Curr Protoc Stem CellBiol. Chapter 1:Unit 1C.2, 2007. It will be understood that cultureconditions can be feeder layer free. It will also be understood that theculture conditions can include the use of matrices such as laminin,Matrigel™, and the like. In some embodiments, methods described in ChenA E, et al., Optimal timing of inner cell mass isolation increases theefficiency of human embryonic stem cell derivation and allows generationof sibling cell lines. Cell Stem Cell. 4(2): 103-6, 2009, are used. Insome embodiments, mitomycin C-inactivated mouse embryonic fibroblastfeeder cells are used for culturing human embryonic stem cells. It willbe understood that in some embodiments other methods of inactivatingfeeder cells may be used such as other compounds or gamma irradiation.

A pluripotent vertebrate cell can be converted to a more naïve state byculturing the cell in the presence of a serine/threonine-protein kinaseB-Raf (BRAF) inhibitor, an epidermal growth factor receptor (EGFR)inhibitor, a vascular endothelial growth factor 1 (VEGFR1) inhibitor, ora fibroblast growth factor receptor 1 (FGFR1) inhibitor; and maintainingthe cell in culture under conditions suitable and a time sufficient tochange the pluripotency state of the vertebrate cell to a more naïvestate.

The cell is cultured and maintained in a culture medium under suitableconditions until the pluripotency state of the cell is converted to amore naïve state. Conditions suitable for conversion of the pluripotentstate of the cell to a more naïve state include the presence of one ormore kinase inhibitors in the culture medium. Thus, in some embodiments,the culture medium contains a serine/threonine-protein kinase B-Raf(BRAF) inhibitor, an epidermal growth factor receptor (EGFR) inhibitor,a vascular endothelial growth factor 1 (VEGFR1) inhibitor, or afibroblast growth factor receptor 1 (FGFR1) inhibitor. In someembodiments, the culture medium further contains a mitogen-activatedprotein kinase kinase (MEK) inhibitor. In some embodiments, the culturemedium further contains a ROCK inhibitor and/or an Src inhibitor. Insome embodiments, the culture medium further contains a glycogensynthase kinase 3 (GSK3) inhibitor, a rho-associated protein kinase(ROCK) inhibitor, and/or a proto-oncogene tyrosine-protein kinase (Src)inhibitor. In some embodiments, the culture medium contains a BRAFinhibitor, a MEK inhibitor, a GSK3 inhibitor, a ROCK inhibitor, and anSrc inhibitor. In some embodiments, the culture medium further containsfibroblast growth factor and/or Activin A. In some embodiments, theculture medium contains one or more of the compounds described herein.In some embodiments, the culture medium contains a BRAF inhibitor, a MEKinhibitor, a ROCK inhibitor, and an Src inhibitor. In some embodiments,the culture medium does not contain a GSK3 inhibitor.

The concentration of the kinase inhibitors used in the culture mediumwill depend on the amount of culture medium being generated. In someembodiments, 0.1 μM, 0.2 μM, 0.3 μM, 0.4 μM, 0.5 μM, 1 μM, 1.5 μM, 2.0μM, 2.5 μM, 3.0 μM, 3.5 μM, 4.0 μM, 4.5 μM, 5.0 μM, 5.5 μM, 6.0 μM, 6.5μM, 7.0 μM, 8.0 μM, 8.5 μM, 9.0 μM, 9.5 μM, 10.5 μM, 11.0 μM, 12.0 μM13.0 μM, 14.0 μM, or 15.0 μM of one or more kinase inhibitors areincluded in about 500 mL of culture medium. In some embodiments, 0.1-1.0μM of BRAF inhibitor is used in about 500 ml of culture medium. In someembodiments, 0.5-1.5 μM of MEK inhibitor is used in about 500 ml ofculture medium. In some embodiments, 0.5-1.5 μM of Src inhibitor is usedin about 500 ml of culture medium. In some embodiments, 5.0-15 μM ofROCK inhibitor is used in about 500 ml of culture medium. In someembodiments, 1.0-5.0 μM of GSK3 inhibitor is used in about 500 ml ofculture medium.

In some embodiments, the concentration of GSK3 inhibitor is 1 nM to 10nM. In some embodiments, the concentration of GSK3 inhibitor is 10 nM to0.1 μM. In some embodiments, the concentration of GSK3 inhibitor is 0.1μM to 0.2 μM. In some embodiments, the concentration of GSK3 inhibitoris 0.2 μM to 0.5 μM. In some embodiments, the concentration of GSK3inhibitor is 0.5 μM to 1 μM. In some embodiments, the culture mediumcontains a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, and an Srcinhibitor, and further contains a GSK3 inhibitor in concentration rangelisted above in this paragraph.

In some embodiments, if a different kinase inhibitor that targets thesame kinase is used, such kinase inhibitor may be used at aconcentration that provides an approximately equivalent effect.

In some embodiments, conditions suitable for conversion of thepluripotent state of the cell to a more naïve state include, in additionto the one or more kinase inhibitors described herein, the addition ofone or more of the following components to the culture medium: N2supplement, B27 supplement, human LIF, glutamine, nonessential aminoacids, β-mercaptoethanol, penicillin-streptomycin, and/or albumin (e.g.,BSA or human albumin). In some embodiments, the culture media compriseDMEM/F12 or Neurobasal. In some embodiments, the culture media compriseDMEM/F12 and Neurobasal in a suitable ratio (e.g., 10:90 to 90:10 (e.g.,25:75, 50:50, and 75:25) DMEM/F12: Neurobasal). In some embodiments, theculture media comprising DMEM/F12 and/or Neurobasal further comprise N2supplement, B27 supplement, human LIF, glutamine, nonessential aminoacids, β-mercaptoethanol, penicillin-streptomycin, and/or albumin (e.g.,BSA or human albumin). In some embodiments, the culture medium furthercomprises fibroblast growth factor 2 (FGF2) and 1%, 0.8%, 0.6%, 0.4%,0.2%, or 0.1% KSR. In some embodiment, the cells are maintained onmitomycin C-inactivated mouse embryonic fibroblast feeder cells. In someembodiments, conditions suitable for conversion of the pluripotent stateof the cell to a more naïve state include growing the cells underphysiological oxygen conditions, i.e., about 5% O₂. In some embodiments,the cells are grown under between about 1% O₂ and about 5% O₂. In someembodiments, the cells are grown under between about 2% O₂ and about 5%O₂. In some embodiments, the cells are grown under between about 5% O₂and about 10% O₂. In some embodiments, the cells are grown under betweenabout 10% O₂ and about 20% O₂. In some embodiments, the cells are grownunder 1% O₂, 2% O₂, 3% O₂, 4% O₂, 5% O₂, 6% O₂, 7% O₂, 8% O₂, 9% O₂, 10%O₂, 11% O₂, 12% O₂, 13% O₂, 14% O₂, 15% O₂, 16% O₂, 17% O₂, 18% O₂, 19%O₂, 20% O₂, 21% O₂, 22% O₂, 23% O₂, 24% O₂, or 25% O₂.

Some aspects of the invention relate to a cell culture mediumcomprising: a basal medium; and a serine/threonine-protein kinase B-Raf(BRAF) inhibitor, an epidermal growth factor receptor (EGFR) inhibitor,a vascular endothelial growth factor 1 (VEGFR1) inhibitor, or afibroblast growth factor receptor 1 (FGFR1) inhibitor. In someembodiments, the cell culture medium further comprises mitogen-activatedprotein kinase kinase (MEK) inhibitor. In some embodiments, the cellculture medium further comprises a glycogen synthase kinase 3 (GSK3)inhibitor, a rho-associated protein kinase (ROCK) inhibitor, and/or aproto-oncogene tyrosine-protein kinase (Src) inhibitor. In someembodiments, the culture medium further contains a ROCK inhibitor and/oran Src inhibitor. In some embodiments, the cell culture medium comprisesa BRAF inhibitor, a MEK inhibitor, and a GSK3 inhibitor. In someembodiments, the cell culture medium comprises a BRAF inhibitor, a MEKinhibitor, a GSK3 inhibitor, and a ROCK inhibitor. In some embodiments,the cell culture medium comprises a BRAF inhibitor, a MEK inhibitor, aGSK3 inhibitor, a ROCK inhibitor, and an Src inhibitor. In someembodiments, the culture medium contains an inhibitor of a receptortyrosine kinase, a MEK inhibitor, and a GSK3 inhibitor. In someembodiments, the culture medium contains a BRAF inhibitor, a MEKinhibitor, a GSK3 inhibitor, and an Src inhibitor. In some embodiments,any of the culture media contains a VEGRF1 inhibitor, an EGFR inhibitor,a FGFR1 inhibitor, or a combination thereof, instead of, or in additionto, a BRAF inhibitor. In some embodiments, the culture medium contains aBRAF inhibitor and a MEK inhibitor. In some embodiments, the culturemedium contains a BRAF inhibitor, a MEK inhibitor, and a ROCK inhibitor.In some embodiments, the culture medium contains a BRAF inhibitor, a MEKinhibitor, a ROCK inhibitor, and an Src inhibitor. In some embodiments,the culture medium contains an inhibitor of a receptor tyrosine kinaseand a MEK inhibitor. In some embodiments, the culture medium contains aBRAF inhibitor, a MEK inhibitor, and an Src inhibitor. In someembodiments, the culture medium does not contain a GSK3 inhibitor.

In some embodiments, the concentration of GSK3 inhibitor is 1 nM to 10nM. In some embodiments, the concentration of GSK3 inhibitor is 10 nM to0.1 M. In some embodiments, the concentration of GSK3 inhibitor is 0.1 Mto 0.2 M. In some embodiments, the concentration of GSK3 inhibitor is0.2 M to 0.5 M. In some embodiments, the concentration of GSK3 inhibitoris 0.5 M to 1 M. In some embodiments, the culture medium contains a BRAFinhibitor, a MEK inhibitor, a ROCK inhibitor, and an Src inhibitor, andfurther contains a GSK3 inhibitor in concentration range listed above inthis paragraph. In some embodiments, the culture medium contains a BRAFinhibitor and a MEK inhibitor, and further contains a GSK3 inhibitor inconcentration range listed above in this paragraph. In some embodiments,the culture medium contains a BRAF inhibitor, a MEK inhibitor, and aROCK inhibitor and further contains a GSK3 inhibitor in concentrationrange listed above in this paragraph. In some embodiments, the culturemedium contains a receptor tyrosine kinase and a MEK inhibitor, andfurther contains a GSK3 inhibitor in concentration range listed above inthis paragraph. In some embodiments, the culture medium contains BRAFinhibitor, a MEK inhibitor, and an Src inhibitor, and further contains aGSK3 inhibitor in concentration range listed above in this paragraph.

As used herein, a “basal medium” is typically an unsupplemented medium(e.g., Eagle's minimal essential medium (EMEM); Dulbecco's modifiedEagle's medium (DMEM)). As will be appreciated by those of skill in theart, a basal medium can comprises a variety of components such as one ormore amino acids (e.g., non-essential amino acids, essential aminoacids), salts (e.g., calcium chloride, potassium chloride, magnesiumsulfate, sodium chloride, and monosodium phosphate), sugars (e.g.,glucose), and vitamins (e.g., folic acid, nicotinamide, riboflavin,B12), iron and pH indicators (e.g., phenol red). The basal medium canfurther comprise proteins (e.g., albumin), hormones (e.g., insulin),glycoproteins (e.g., transferrin), minerals (e.g., selenium), serum(e.g., fetal bovine serum), antibiotics, antimycotics andglycosaminoglycans.

In some embodiments, the basal medium is serum-free medium. In someembodiments, the basal medium comprises one or more supplements, suchas, but not limited to, supplements such as B27 and/or N2. In someembodiments, the basal medium is supplemented with one or more of thefollowing: DMEM/F12, Neurobasal, N2 supplement, 10 mL B27 supplement,human LIF, glutamine, nonessential amino acids, β-mercaptoethanol,penicillin-streptomycin, and/or BSA (Sigma). In some embodiments, thesupplemented basal cell culture medium further comprises fibroblastgrowth factor 2 (FGF2) and 1%, 0.8%, 0.6%, 0.4%, 0.2%, or 0.1% KSR.

The pluripotent vertebrate cell is maintained in culture for a timesufficient to change the pluripotency state of the cell to a more naïvestate. In some embodiments, time sufficient to change the pluripotencystate of the cell to a more naïve state is at least 1 day, at least 2days, at least 3 days, at least 4 days, at least 5 days, at least 6days, at least 7 days, at least 8 days, at least 9 days, at least 10days, at least 11 days, at least 12 days, at least 13 days, at least 14days, at least 15 days, at least 16 days, at least 17 days, at least 18days, at least 19 days, or at least 20 days. In some embodiments, timesufficient to change the pluripotency state of the cell to a more naïvestate is between 1-5 days, 1-10 days, 1-15 days, 1-20 days, 5-10 days,5-15 days, 5-20 days, 10-15 days, 10-20 days, or 15-20 days. In someembodiments, time sufficient to change the pluripotency state of thecell to a more naïve state is at least about 5 days (e.g., about 10days). In some embodiments, the culture medium is replenished asrequired during this time. In some embodiments, the cell is maintainedin culture until the cell has at least one property which is similar tothe corresponding property of mouse embryonic stem cells. In someembodiments, the at least one property which is similar to thecorresponding property of mouse embryonic stem cells is the utilizationof the distal OCT4 enhancer element. An important molecular signature ofnaïve pluripotency in the mouse system is the use of the distal enhancer(DE) of OCT4. Thus, in some embodiments, the cell is maintained inculture until the cell uses the distal Oct4 enhancer element for OCT4expression. In some embodiments, the cell is maintained in culture untilthe cell uses the endogenous distal Oct4 enhancer element for OCT4expression. In some embodiments, the cell is maintained in culture untilthe cell has enhanced utilization of the distal Oct4 enhancer elementfor OCT4 expression as compared to the cell prior to theculturing/maintenance period. In some embodiments, the cell ismaintained in culture until the cell has enhanced utilization of theendogenous distal Oct4 enhancer element for OCT4 expression as comparedto the cell prior to the culturing/maintenance period. The utilizationof the distal OCT4 enhancer element can be tested using the OCT4-ΔPE-GFPreporter system described in the Examples below.

In some embodiments, the at least one property which is similar to thecorresponding property of mouse embryonic stem cells is colonymorphology. Naïve pluripotent cells that correspond to the more immature“ground state” of pluripotency, exhibit a dome-like colony morphology.Thus, in certain embodiments, the cell is maintained in culture until itexhibits a dome-like colony morphology.

In some embodiments, the at least one property which is similar to thecorresponding property of mouse embryonic stem cells is gene expressionprofile. The cell is maintained in culture until it has a global geneexpression profile which clusters with naïve mouse ESCs as opposed tomouse EpiSCs and/or less naïve human ESCs. In some embodiments, the geneexpression profile includes markers of ground state pluripotency, suchas, but not limited to, NANOG, OCT4, DPPA5, DPPA3 (also known asSTELLA), KLF4, KLF5, TFCP2L1, and/or REX1. In some embodiments, the cellis maintained in culture until it exhibits a gene expression profilesimilar to that shown in FIG. 6.

Some aspects of the invention relate to a method for culturingvertebrate cells, the method comprising growing vertebrate cells in acell culture medium comprising a serine/threonine-protein kinase B-Raf(BRAF) inhibitor, an epidermal growth factor receptor (EGFR) inhibitor,a vascular endothelial growth factor 1 (VEGFR1) inhibitor, or afibroblast growth factor receptor 1 (FGFR1) inhibitor. The vertebratecells can be naïve pluripotent cells, human pluripotent cells, or naïvehuman pluripotent cells. In some embodiments, the cell culture mediumfurther comprises mitogen-activated protein kinase kinase (MEK)inhibitor. In some embodiments, the cell culture medium furthercomprises a glycogen synthase kinase 3 (GSK3) inhibitor, arho-associated protein kinase (ROCK) inhibitor, and/or a proto-oncogenetyrosine-protein kinase (Src) inhibitor. In some embodiments, theculture medium further contains a ROCK inhibitor and/or an Srcinhibitor. In some embodiments, the cell culture medium comprises a BRAFinhibitor, a MEK inhibitor, and a GSK3 inhibitor. In some embodiments,the cell culture medium comprises a BRAF inhibitor, a MEK inhibitor, aGSK3 inhibitor, and a ROCK inhibitor. In some embodiments, the cellculture medium comprises a BRAF inhibitor, a MEK inhibitor, a GSK3inhibitor, a ROCK inhibitor, and an Src inhibitor. In some embodiments,the culture medium contains a BRAF inhibitor and a MEK inhibitor. Insome embodiments, the culture medium contains a BRAF inhibitor, a MEKinhibitor, and a ROCK inhibitor. In some embodiments, the culture mediumcontains a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, and an Srcinhibitor. In some embodiments, the culture medium does not contain aGSK3 inhibitor.

In some embodiments, the concentration of GSK3 inhibitor is 1 nM to 10nM. In some embodiments, the concentration of GSK3 inhibitor is 10 nM to0.1 μM. In some embodiments, the concentration of GSK3 inhibitor is 0.1μM to 0.2 μM. In some embodiments, the concentration of GSK3 inhibitoris 0.2 μM to 0.5 μM. In some embodiments, the concentration of GSK3inhibitor is 0.5 μM to 1 μM. In some embodiments, the culture mediumcontains a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, and an Srcinhibitor, and further contains a GSK3 inhibitor in concentration rangelisted above in this paragraph. In some embodiments, the culture mediumcontains a BRAF inhibitor and a MEK inhibitor, and further contains aGSK3 inhibitor in concentration range listed above in this paragraph. Insome embodiments, the culture medium contains a BRAF inhibitor, a MEKinhibitor, and a ROCK inhibitor and further contains a GSK3 inhibitor inconcentration range listed above in this paragraph.

The cell culture medium may be a basal medium. As used herein, a “basalmedium” is typically an unsupplemented medium (e.g., Eagle's minimalessential medium (EMEM); Dulbecco's modified Eagle's medium (DMEM)). Aswill be appreciated by those of skill in the art, a basal medium cancomprises a variety of components such as one or more amino acids (e.g.,non-essential amino acids, essential amino acids), salts (e.g., calciumchloride, potassium chloride, magnesium sulfate, sodium chloride, andmonosodium phosphate), sugars (e.g., glucose), and vitamins (e.g., folicacid, nicotinamide, riboflavin, B12), iron and pH indicators (e.g.,phenol red). The basal medium can further comprise proteins (e.g.,albumin), hormones (e.g., insulin), glycoproteins (e.g., transferrin),minerals (e.g., selenium), serum (e.g., fetal bovine serum),antibiotics, antimycotics and glycosaminoglycans.

In some embodiments, the basal medium is serum-free medium. In someembodiments, the basal medium is supplemented with one or more of thefollowing: DMEM/F12, Neurobasal, N2 supplement, 10 mL B27 supplement,human LIF, glutamine, nonessential amino acids, β-mercaptoethanol,penicillin-streptomycin, and/or BSA (Sigma). In some embodiments, thesupplemented basal medium further comprises fibroblast growth factor 2(FGF2) and 1%, 0.8%, 0.6%, 0.4%, 0.2%, or 0.1% KSR.

Some aspects of the disclosure relate to naïve pluripotent vertebratecells produced by the methods described herein. Some aspects of thedisclosure relate to compositions comprising the naïve pluripotentvertebrate cells produced by the methods described herein. Thedisclosure provides pluripotent cells, cell lines, and cell clonesderived or cultured using the methods and/or compositions describedherein. The invention further provides cell cultures, wherein at leastsome of the cells in the cell culture are derived or cultured using themethods and/or compositions described herein. Some aspects of theinvention relate to a naïve pluripotent vertebrate cell line, e.g., anaïve pluripotent human cell line, produced by the methods describedherein. In some aspects, a naïve pluripotent vertebrate cell isprovided, wherein the cell uses the distal Oct4 enhancer element forOCT4 expression. In some embodiments, the cell primarily uses theendogenous distal OCT4 enhancer element, e.g., the ratio of the cell'suse of endogenous distal OCT4 enhancer to the cell's use of endogenousproximal OCT4 enhancer is at least about 1:1 (e.g., about 2:1, about3:1, about 5:1, or about 10:1). In some aspects, a naïve pluripotentvertebrate cell, e.g., a naïve pluripotent human cell, is provided,wherein the cell has a global gene expression profile which clusterswith naïve mouse ESCs as opposed to stem cell lines derived from mouseepiblast (EpiSCs) and/or less naïve human ESCs. In some embodiments,these cells are produced by methods described herein. In some aspects,provided herein are conventional pluripotent vertebrate cells (e.g.,conventional pluripotent human cells) with a deletion of the proximalOCT4 enhancer element (or other disabling mutation). In some aspects,provided herein are methods of using the conventional pluripotentvertebrate cells (e.g., conventional pluripotent human cells) with adeletion of the proximal OCT4 enhancer element (or other disablingmutation), examples of such methods including methods of identifyingcompounds that induce a naïve pluripotent state.

In some embodiments, at least 80% or at least 90% of the pluripotentstem cells of a colony, cell line, or cell culture express one or moremarker(s), e.g., a set of markers, indicative of pluripotency, e.g., aground state of pluripotency. In some embodiments at least 91%, at least92%, at least 93%, at least 94%, at least 95%, at least 96%, at least97%, at least 98%, at least 99%, or more of the cells of a colony, cellline, or cell culture express the marker(s).

Some aspects of the invention provide a kit for culturing vertebratecells. The kit comprises a serine/threonine-protein kinase B-Raf (BRAF)inhibitor, an epidermal growth factor receptor (EGFR) inhibitor, avascular endothelial growth factor 1 (VEGFR1) inhibitor, or a fibroblastgrowth factor receptor 1 (FGFR1) inhibitor. In some embodiments, the kitalso comprises instructions for culturing vertebrate cells.

In some embodiments, the kit further comprises mitogen-activated proteinkinase kinase (MEK) inhibitor. In some embodiments, the kit furthercomprises a glycogen synthase kinase 3 (GSK3) inhibitor, arho-associated protein kinase (ROCK) inhibitor, and/or a proto-oncogenetyrosine-protein kinase (Src) inhibitor. In some embodiments, the kitfurther comprises a ROCK inhibitor and/or an Src inhibitor. In someembodiments, the kit comprises a BRAF inhibitor, a MEK inhibitor, and aGSK3 inhibitor. In some embodiments, the kit comprises a BRAF inhibitor,a MEK inhibitor, a GSK3 inhibitor, and a ROCK inhibitor. In someembodiments, the kit comprises a BRAF inhibitor, a MEK inhibitor, a GSK3inhibitor, a ROCK inhibitor, and an Src inhibitor. In some embodiments,the kit comprises a BRAF inhibitor and a MEK inhibitor. In someembodiments, the kit comprises a BRAF inhibitor, a MEK inhibitor, and aROCK inhibitor. In some embodiments, the kit comprises a BRAF inhibitor,a MEK inhibitor, a ROCK inhibitor, and an Src inhibitor. In someembodiments, the kit does not comprise a GSK inhibitor. In someembodiments, the kit further comprises cell culture medium. Thevertebrate cells can be naïve pluripotent cells, human pluripotentcells, or naïve human pluripotent cells. The compositions or contents ofthe kits can be provided in one or more containers (e.g., compounds thatare compatible can be provided together in the same container). Thecomponents of the kit may be sterile.

Some aspects of the invention relate to a kit for preparing a cellculture medium. The kit comprises a serine/threonine-protein kinaseB-Raf (BRAF) inhibitor, an epidermal growth factor receptor (EGFR)inhibitor, a vascular endothelial growth factor 1 (VEGFR1) inhibitor, ora fibroblast growth factor receptor 1 (FGFR1) inhibitor; andinstructions for preparing a cell culture medium. In some embodiments,the kit further comprises mitogen-activated protein kinase kinase (MEK)inhibitor. In some embodiments, the kit further comprises a glycogensynthase kinase 3 (GSK3) inhibitor, a rho-associated protein kinase(ROCK) inhibitor, and/or a proto-oncogene tyrosine-protein kinase (Src)inhibitor. In some embodiments, the kit further comprises a ROCKinhibitor and/or an Src inhibitor. In some embodiments, the kitcomprises a BRAF inhibitor, a MEK inhibitor, and a GSK3 inhibitor. Insome embodiments, the kit comprises a BRAF inhibitor, a MEK inhibitor, aGSK3 inhibitor, and a ROCK inhibitor. In some embodiments, the kitcomprises a BRAF inhibitor, a MEK inhibitor, a GSK3 inhibitor, a ROCKinhibitor, and an Src inhibitor. In some embodiments, the kit comprisesa BRAF inhibitor and a MEK inhibitor.

In some embodiments, the kit comprises a BRAF inhibitor, a MEKinhibitor, and a ROCK inhibitor. In some embodiments, the kit comprisesa BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, and an Srcinhibitor. In some embodiments, the kit does not comprise a GSK3inhibitor.

In some embodiments, the kit further comprises a basal medium. As usedherein, a “basal medium” is typically an unsupplemented medium (e.g.,Eagle's minimal essential medium (EMEM); Dulbecco's modified Eagle'smedium (DMEM)). As will be appreciated by those of skill in the art, abasal medium can comprises a variety of components such as one or moreamino acids (e.g., non-essential amino acids, essential amino acids),salts (e.g., calcium chloride, potassium chloride, magnesium sulfate,sodium chloride, and monosodium phosphate), sugars (e.g., glucose), andvitamins (e.g., folic acid, nicotinamide, riboflavin, B12), iron and pHindicators (e.g., phenol red). The basal medium can further compriseproteins (e.g., albumin), hormones (e.g., insulin), glycoproteins (e.g.,transferrin), minerals (e.g., selenium), serum (e.g., fetal bovineserum), antibiotics, antimycotics and glycosaminoglycans.

In some embodiments, the basal medium is serum-free medium. In someembodiments, the basal medium comprises one or more supplements, suchas, but not limited to, supplements such as B27 and/or N2. In someembodiments, the basal medium is supplemented with one or more of thefollowing: DMEM/F12, Neurobasal, N2 supplement, 10 mL B27 supplement,human LIF, glutamine, nonessential amino acids, β-mercaptoethanol,penicillin-streptomycin, and/or BSA (Sigma). In some embodiments, thesupplemented basal cell culture medium further comprises fibroblastgrowth factor 2 (FGF2) and 1%, 0.8%, 0.6%, 0.4%, 0.2%, or 0.1% KSR.

Some aspects of the invention provide a kit for changing thepluripotency state of a vertebrate cell to a more naïve state. The kitcomprises a pluripotent vertebrate cell; and cell culture mediumcomprising a serine/threonine-protein kinase B-Raf (BRAF) inhibitor, anepidermal growth factor receptor (EGFR) inhibitor, a vascularendothelial growth factor 1 (VEGFR1) inhibitor, or a fibroblast growthfactor receptor 1 (FGFR1) inhibitor. In some embodiments, the cellculture medium further comprises mitogen-activated protein kinase kinase(MEK) inhibitor. In some embodiments, the cell culture medium furthercomprises a glycogen synthase kinase 3 (GSK3) inhibitor, arho-associated protein kinase (ROCK) inhibitor, and/or a proto-oncogenetyrosine-protein kinase (Src) inhibitor. In some embodiments, the cellculture medium further comprises a ROCK inhibitor and/or an Srcinhibitor. In some embodiments, the cell culture medium comprises a BRAFinhibitor, a MEK inhibitor, and a GSK3 inhibitor. In some embodiments,the cell culture medium comprises a BRAF inhibitor, a MEK inhibitor, aGSK3 inhibitor, and a ROCK inhibitor. In some embodiments, the cellculture medium comprises a BRAF inhibitor, a MEK inhibitor, a GSK3inhibitor, a ROCK inhibitor, and an Src inhibitor. In some embodiments,the cell culture medium comprises a BRAF inhibitor and a MEK inhibitor.In some embodiments, the cell culture medium comprises a BRAF inhibitor,a MEK inhibitor, and a ROCK inhibitor. In some embodiments, the cellculture medium comprises a BRAF inhibitor, a MEK inhibitor, a ROCKinhibitor, and an Src inhibitor. In some embodiments, the cell culturemedium does not comprise a GSK3 inhibitor.

Gene expression profiling may be used to assess the pluripotency stateof a cell or population of cells. Primed or conventional pluripotentcells, such as embryonic stem cells, and multipotent cells (i.e., adultstem cells) exhibit a distinct pattern of global gene expression ascompared to more naïve pluripotent cells (i.e., the more immature“ground state” of pluripotency). Thus, one may assess DNA methylation,gene expression, and/or epigenetic state of cellular DNA, and/ordevelopmental potential of the cells, e.g., as described in Wernig etal., Nature, 448:318-24, 2007. Other methods of assessing pluripotencystate include epigenetic analysis, e.g., analysis of DNA methylationstate (Nazor et al. Cell Stem Cell. 2012 May 4; 10(5):620-34).

In some embodiments a pluripotent cell described herein is used toprepare a composition for cell therapy to be administered to avertebrate subject, e.g., a non-human animal, or a human. In someembodiments, a pluripotent cell derived or cultured according to thesystems and methods described herein is used to prepare a compositionfor cell therapy to be administered to a vertebrate subject, e.g., anon-human animal, or a human. In some embodiments, the compositioncomprises autologous cells. In other embodiments, the compositioncomprises non-autologous cells. In some embodiments, the cells aregenetically matched to an individual.

The naïve pluripotent vertebrate stem cells may be used to treat anumber of life-threatening diseases and disorders. Examples include, butare not limited to, cancers (such as, but not limited to, acuteleukemia, chronic leukemia, high-risk solid tumors, Hodgkin &Non-Hodgkin Lymphoma, myelodysplastic syndromes), blood disorders (suchas, but not limited to, aplastic anemia, beta thalassemia,Diamond-Blackfan Anemia, Fanconi Anemia, Sickle Cell Disease), immunedisorders (such as, but not limited to, chronic granulomatous disease,hystiocytic disorders, leukocyte adhesion deficiency, severe combinedimmunodeficiency diseases, Wiskott-Aldrich Syndrome), and metabolicdisorders (such as, but not limited to, Krabbe Disease, Hurler Syndrome,Metachromatic Leukodystrophy, and Sanfilippo Syndrome). The naïvepluripotent vertebrate stem cells produced by the methods describedherein are administered to a subject having or suspected of having adisease or disorder that can be treated using stem cell therapy. Severalapproaches may be used for the introduction of the naïve pluripotentvertebrate stem cells stem cells into the subject, including but notlimited to, catheter-mediated delivery I.V. (e.g., endovascularcatheter), direct injection into a target site, intravenous injection,intraperitoneal injection, parenteral injection, subcutaneous injection,intramuscular injection, and/or intracardiac injection.

The compositions and methods of the disclosure may be applied to deriveor culture naïve pluripotent cells from non-human animals including butnot limited to, dogs, cats, horses, sheep, goats, cows, and/or rodents(such as rats, rabbits, hamsters, guinea pigs).

The disclosure may be applied to derive or culture naïve pluripotentcells from primates, e.g., non-human primates, or humans. In manyembodiments, the vertebrate is a mammal. In some embodiments the mammalis a bovine, ovine, caprine, equine, canine, or feline. It is alsoenvisioned that compositions and methods of the invention may be used toderive naïve pluripotent cells, e.g., ES cells or iPS cells fromnon-mammalian vertebrates, e.g., zebrafish or other non-mammalianorganisms of interest such as birds. In some embodiments, if the speciesis one from which ES or iPS cells have not heretofore been derived,techniques and culture conditions can be adapted from standardtechniques used in other species, e.g., related species.

Systems, compositions, and methods of the invention can be applied inthe derivation or culture of naïve pluripotent cells derived from cellsobtained from any of a variety of sources. For example, cells obtainedfrom the inner cell mass (ICM) or epiblast can be used to derive naïveES cells. In some embodiments, the systems, compositions, and methodsare applied to derive naïve pluripotent stem cells from blastomeres,e.g., blastomeres isolated from a morula or from a 4-8 cell stageembryo. In some embodiments, the compositions and methods are applied toderive naïve pluripotent stem cells from germ cells. In some embodimentsthe compositions and methods are used to derive naïve pluripotent cellsusing parthenogenesis or SCNT. In some embodiments the methods areapplied to derive or culture naïve induced pluripotent stem (iPS) cells.Methods for generating iPS cells are well-known in the art (see, forexample, WO2013159103, WO 2013177133, and U.S. Pat. No. 8,748,179).iPSCs are typically derived by introducing a specific set ofpluripotency-associated genes, or “reprogramming factors”, into a givencell type. The original set of reprogramming factors are the genes Oct4(Pou5fl), Sox2, cMyc, and Klf4. While this combination is mostconventional in producing iPSCs, each of the factors can be functionallyreplaced by related transcription factors, miRNAs, small molecules, oreven non-related genes such as lineage specifiers. In some embodiments,somatic cells used to generate iPS cells include, but are not limitedto, fibroblasts, keratinocytes, immune system cells, and epithelialcells. In some embodiments, iPS cells are generated without genomicmodification. In some embodiments, iPS cells are free of exogenouslyintroduced DNA. For example, they may be generated using syntheticmodified mRNA, small molecules, or a combination thereof. For example,iPS cells may be generated using the methods described in Mandal, P K &Rossi, D J, Nature Protocols 8, 568-582 (2013). In some embodiments, iPScells are generated using episomal expression of one or more of thereprogramming factors. After the iPS cells are generated, the episome(s)may be lost resulting in cells free of exogenously introduced DNA.

The invention contemplates a variety of uses for the pluripotent cells,cell lines derived, cultured, or generated as described herein. Ingeneral, pluripotent cells may be used for any purpose contemplated inthe art for use of pluripotent cells, e.g., ES or iPS cells. See, e.g.,international PCT applications, no. PCT/US2013/050077 (WO 2014/011881)and PCT/US2001/006912 (WO 2001/066697).

In some embodiments a pluripotent cell derived or cultured according tothe invention is used to produce one or more differentiated cells. Suchcells are considered to be an aspect of the disclosure. The cells couldbe, e.g., multipotent stem cells or fully differentiated cells. Thecells may be, e.g., hematopoietic cells (e.g., of the myeloid orerythroid lineage), neural cells (e.g., neural precursors, neurons orglial cells), myoblasts, myocytes, cardiomyocytes, melanoblasts,keratinocytes, chondroblasts, chondrocytes, osteoblasts, osteoclasts,pancreatic beta cells, retinal cells, endothelial cells, etc. Protocolsknown in the art for differentiating cells into cells of a desired typemay be used (see, for example, US 20130273651). In some embodiments apluripotent cell may be differentiated to a cell type of interest exvivo, e.g., before being administered to a subject. For example, apluripotent cell may be differentiated to produce cells of a cell typethat is affected by a disease or that may be useful in treating adisease for which the subject is in need of treatment. In someembodiments cells are used to generate a tissue or organ in vitro or tosupplement a tissue or organ in vivo.

The disclosure also provides methods of producing non-human vertebrates,e.g., non-human mammals, which can be genetically modified ornon-genetically modified, using the pluripotent cells of the disclosure.Such non-human vertebrates are aspects of the disclosure. In someembodiments the non-human vertebrates are mice. In some embodiments,non-human mammals are produced using methods known in the art forproducing non-human mammals from ES or iPS cells (see, for example, WO2010124290). For example, ES or iPS cells are introduced into ablastocyst of the same species which is transferred to a suitable fostermother (e.g., a pseudopregnant female of the same species) underconditions suitable for production of live offspring. If a diploidblastocyst is used, chimeric offspring may be produced, which aretypically derived in part from the ES cell or iPS cell and in part fromthe blastocyst into which the cell was introduced. Chimeric offspringmay be interbred to generate homozygous animals if the chimericoffspring contain ES-derived contribution to the germ line as known inthe art. In some embodiments, the mice are produced using methods thatdo not require production of chimera or chimeric offspring. In someembodiments, pluripotent ES cells are introduced into tetraploidblastocysts of the same vertebrate species under conditions that resultin production of an embryo (at least one/one or more embryos) and theresulting embryo(s) transferred into an appropriate foster mother, suchas a pseudopregnant female of the same vertebrate species. The resultingfemale is maintained under conditions that result in development of liveoffspring, thereby producing a non-human mammal derived from theintroduced ES cells. See, e.g., U.S. Pat. No. 6,784,336. In someembodiments, the mouse is produced by a method that involveslaser-assisted injection or piezo-injection of ES cells of the inventioninto four- or eight-cell embryos. In some embodiments the mouse isproduced without need to generate a chimera, e.g., using methodsdescribed in international PCT application, no. PCT/EP2003/002180 (WO2003/073843). Another embodiment of the present invention is a method ofproducing a non-human mammalian strain, such as a mouse strain, e.g., agenetically engineered mouse strain, that is derived from a given(single) iPS or ES cell clone of the present disclosure withoutoutcrossing with a wild type partner. See, e.g., U.S. Pat. No.6,784,336. In some embodiments the mice are genetically modified, e.g.,they are derived from an ES or iPS cell that is genetically modified.The invention contemplates interbreeding non-human vertebrates, e.g.,mice, derived from the ES cells or iPS cells with mice of any strain ofinterest, the resulting strains being considered other aspects of theinvention.

A naïve pluripotent cell can be derived from a cell, e.g., a somaticcell, obtained from an individual of interest. The individual can be,e.g., a human suffering from a disease or condition. In some embodimentsthe individual is immunocompatible with an individual suffering from adisease or condition. In some embodiments the disease is aneurodegenerative disease, e.g., Parkinson's disease, Alzheimer'sdisease, or amyotrophic lateral sclerosis. In some embodiments theindividual suffers from diabetes. In some embodiments the individualsuffers from heart failure or a muscle disorder or an enzyme deficiency,sickle cell anemia, hemophilia, a glycogen storage disorder, or cysticfibrosis. In some embodiments the disease is a heritable disease. Insome embodiments the disease is a monogenic disorder. In someembodiments the disease has an autosomal dominant inheritance pattern.In some embodiments the disease has an autosomal recessive inheritancepattern. In some embodiments the disease is sporadic, i.e., there is noevident pattern of inheritance. In some embodiments the individual hassuffered an injury, e.g., traumatic brain injury, spinal cord injury. Insome embodiments the individual is in need of cell therapy.

In some embodiments a naïve pluripotent cell, e.g., a naïve pluripotenthuman cell, is derived from a cell (e.g., a somatic cell) obtained froman individual who harbors a mutation or genetic variation that is knownto cause or suspected of causing a disease (or an immunocompatibledonor). The mutation or genetic variation is corrected ex vivo.Resulting cells or cells derived therefrom are administered to thesubject.

In some embodiments, a naïve pluripotent cell, e.g., a naïve pluripotenthuman cell, may be used to generate a model of a disease, e.g., ananimal model or a cellular model of a disease, e.g., any of the diseasesmentioned herein or other diseases of interest. In some embodiments thenaïve pluripotent cell is derived from a cell, e.g., a somatic cell,obtained from an individual who has the disease. In some embodiments thenaïve pluripotent cell is genetically engineered to harbor a mutation orgenetic variation that is known to cause or suspected of causing thedisease. In some embodiments the naïve pluripotent cell may bedifferentiated to a cell of a cell type that is affected by the disease.

In some embodiments, a naïve pluripotent cell is not geneticallymodified. In some embodiments, the naïve pluripotent cell is devoid ofDNA or genetic alterations (e.g., insertions, substitutions, deletions)introduced by the hand of man. In some embodiments, a naïve pluripotentcell may be genetically modified after it is derived. Any of a varietyof different methods may be employed to genetically modify a naïvepluripotent cell or a cell derived therefrom, e.g., a differentiatedcell. Examples of such methods include, but are not limited to,homologous recombination and transfection (see, for example, WO2013042731, and U.S. Pat. No. 8,637,311). In some embodiments, genomeediting technologies such as, but not limited to zinc fingers, TALENs,CRISPR/Cas systems are used to genetically modify the naïve pluripotentcell.

Compounds

Exemplary compounds that are useful in a composition, system, kit, ormethod described herein are provided below.

Compounds of Formula (A)

In one aspect, the present disclosure provides compounds of Formula (A):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(A1) is a substituted or unsubstituted, saturated or unsaturated, C₁₋₄aliphatic chain, optionally wherein one, two, or three chain atoms ofthe aliphatic chain are independently replaced with —O—, —S—, —NR^(A5)—,—N═, or ═N—, wherein each instance of R^(A5) is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;

L^(A2) is a substituted or unsubstituted, saturated or unsaturated, C₁₋₆aliphatic chain, optionally wherein one, two, or three chain atoms ofthe aliphatic chain are independently replaced with —O—, —S—, —NR^(A6)—,—N═, or ═N—, wherein each instance of R^(A6) is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;

R^(A1) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(A2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

n2 is 0, 1, 2, 3, 4, 5, 6, or 7;

each instance of R^(A3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

n3 is 0, 1, 2, or 3;

each instance of R^(A4) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

n4 is 0, 1, 2, 3, or 4.

In certain embodiments, L^(A1) is a substituted or unsubstituted,saturated or unsaturated, C₂ aliphatic chain, optionally wherein onechain atom of the aliphatic chain is replaced with —O—, —S—, —NR^(A5)—(e.g., —NH—), —N═, or ═N—. In certain embodiments, L^(A1) is—C(═O)NR^(A5)— (e.g., —C(═O)NH—) or —NR^(A5)C(═O)— (e.g., —NHC(═O)—). Incertain embodiments, L^(A1) is a substituted or unsubstituted, saturatedor unsaturated, C₁, C₃, or C₄ aliphatic chain, optionally wherein one ortwo chain atoms of the aliphatic chain are independently replaced with—O—, —S—, —NR^(A5)— (e.g., —NH—), —N═, or ═N—.

In certain embodiments, all instances of R^(A5) are the same. In certainembodiments, two instances of R^(A5) are different from each other. Incertain embodiments, at least one instance of R^(A5) is hydrogen. Incertain embodiments, all instances of R^(A5) are hydrogen. In certainembodiments, at least one instance of R^(A5) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(A5) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, L^(A2) is a substituted or unsubstituted,saturated or unsaturated, C₂ aliphatic chain, optionally wherein onechain atom of the aliphatic chain is replaced with —O—, —S—, —NR^(A6)—(e.g., —NH—), —N═, or ═N—. In certain embodiments, L^(A2) is—CH₂NR^(A6)— (e.g., —CH₂NH—) or —NR^(A6)CH₂— (e.g., —NHCH₂—). In certainembodiments, L^(A2) is a substituted or unsubstituted, saturated orunsaturated, C₁, C₃, or C₄ aliphatic chain, optionally wherein one ortwo chain atoms of the aliphatic chain are independently replaced with—O—, —S—, —NR^(A6)— (e.g., —NH—), —N═, or ═N—.

In certain embodiments, all instances of R^(A6) are the same. In certainembodiments, two instances of R^(A6) are different from each other. Incertain embodiments, at least one instance of R^(A6) is hydrogen. Incertain embodiments, all instances of R^(A6) are hydrogen. In certainembodiments, at least one instance of R^(A6) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(A6) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, R^(A1) is hydrogen. In certain embodiments,R^(A1) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(A1) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, two instances of R^(A2) are the same. In certainembodiments, two instances of R^(A2) are different from each other. Incertain embodiments, at least one instance of R^(A2) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(A2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(A2) is —CH₃. In certain embodiments, at least one instance ofR^(A2) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(A2) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(A2) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(A2) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(A2) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(A2) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(A) is substituted or unsubstituted phenyl. In certainembodiments, at least one instance of R^(A2) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(A2) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(A2) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(A2) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(A2)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(A2) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(A2) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(A2) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(A2) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

Each of Formulae (A) to (L) may independently include one or moresubstituents R^(a). In certain embodiments, all instances of R^(a) arethe same. In certain embodiments, two instances of R^(a) are differentfrom each other. In certain embodiments, at least one instance of R^(a)is H. In certain embodiments, at least one instance of R^(a) issubstituted or unsubstituted acyl (e.g., acetyl). In certainembodiments, at least one instance of R^(a) is substituted orunsubstituted alkyl (e.g., substituted or unsubstituted C₁₋₆ alkyl). Incertain embodiments, at least one instance of R^(a) is —CH₃. In certainembodiments, at least one instance of R^(a) is —CF₃, unsubstitutedethyl, perfluoroethyl, unsubstituted propyl, perfluoropropyl,unsubstituted butyl, or perfluorobutyl. In certain embodiments, at leastone instance of R^(a) is substituted or unsubstituted alkenyl (e.g.,substituted or unsubstituted C₁₋₆ alkenyl). In certain embodiments, atleast one instance of R^(a) is substituted or unsubstituted alkynyl(e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, at least one instance of R^(a) is substituted orunsubstituted carbocyclyl (e.g., substituted or unsubstituted,monocyclic, 3- to 7-membered carbocyclyl). In certain embodiments, atleast one instance of R^(a) is substituted or unsubstituted heterocyclyl(e.g., substituted or unsubstituted, monocyclic, 5- to 6-membedheterocyclyl, wherein one, two, or three atoms in the heterocyclic ringsystem are independently nitrogen, oxygen, or sulfur). In certainembodiments, at least one instance of R^(a) is substituted orunsubstituted aryl (e.g., substituted or unsubstituted, 6- to10-membered aryl). In certain embodiments, at least one instance ofR^(a) is substituted or unsubstituted phenyl. In certain embodiments, atleast one instance of R^(a) is substituted or unsubstituted heteroaryl(e.g., substituted or unsubstituted, monocyclic, 5- to 6-membedheteroaryl, wherein one, two, three, or four atoms in the heteroarylring system are independently nitrogen, oxygen, or sulfur). In certainembodiments, at least one instance of R^(a) is a nitrogen protectinggroup (e.g., Bn, Boc, Cbz, Fmoc, trifluoroacetyl, triphenylmethyl,acetyl, or Ts) when attached to a nitrogen atom. In certain embodiments,R^(a) is an oxygen protecting group (e.g., silyl, TBDPS, TBDMS, TIPS,TES, TMS, MOM, THP, t-Bu, Bn, allyl, acetyl, pivaloyl, or benzoyl) whenattached to an oxygen atom. In certain embodiments, R^(a) is a sulfurprotecting group (e.g., acetamidomethyl, t-Bu, 3-nitro-2-pyridinesulfenyl, 2-pyridine-sulfenyl, or triphenylmethyl) when attached to asulfur atom. In certain embodiments, two instances of R^(a) are joinedto form a substituted or unsubstituted heterocyclic ring (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclicring, wherein one, two, or three atoms in the heterocyclic ring systemare independently nitrogen, oxygen, or sulfur). In certain embodiments,two instances of R^(a) are joined to form a substituted or unsubstitutedheteroaryl ring (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heteroaryl ring, wherein one, two, three, or four atoms in theheteroaryl ring system are independently nitrogen, oxygen, or sulfur).

In certain embodiments, n2 is 0. In certain embodiments, n2 is 1. Incertain embodiments, n2 is 2. In certain embodiments, n2 is 3. Incertain embodiments, n2 is 4. In certain embodiments, n2 is 5. Incertain embodiments, n2 is 6. In certain embodiments, n2 is 7.

In certain embodiments, all instances of R^(A3) are the same. In certainembodiments, two instances of R^(A3) are different from each other. Incertain embodiments, at least one instance of R^(A3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(A3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(A3) is —CH₃. In certain embodiments, at least one instance ofR^(A3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(A3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(A3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(A3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(A3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(A3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(A3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(A3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(A3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(A3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(A3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(A3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(A3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(A3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(A3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(A3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, n3 is 0. In certain embodiments, n3 is 1. Incertain embodiments, n3 is 2. In certain embodiments, n3 is 3.

In certain embodiments, all instances of R^(A4) are the same. In certainembodiments, two instances of R^(A4) are different from each other. Incertain embodiments, at least one instance of R^(A4) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(A4) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(A4) is —CH₃. In certain embodiments, at least one instance ofR^(A4) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(A4) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(A4) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(A4) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(A4) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(A4) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(A4) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(A4) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(A4) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(A4) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(A4) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(A4)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(A4) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(A4) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(A4) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(A4) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, n4 is 0. In certain embodiments, n4 is 1. Incertain embodiments, n4 is 2. In certain embodiments, n4 is 3. Incertain embodiments, n4 is 4.

In certain embodiments, the compound of Formula (A) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (A) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (A) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (A) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (A) is not AMG706, or apharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (A), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are VEGFR inhibitors (e.g., VEGFR1 inhibitors).

Compounds of Formula (B)

In another aspect, the present disclosure provides compounds of Formula(B):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(B1) is —N(R^(B4))C(═O)—, —C(═O)N(R^(B4))—, or—N(R^(B4))C(═O)N(R^(B4))—, wherein each instance of R^(B4) isindependently hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

-   -   when L^(B1) is —N(R^(B4))C(═O)— or —C(═O)N(R^(B4))—, L^(B1) is        directly attached to the carbon atom labeled with 3 or 5;    -   when L^(B1) is —N(R^(B4))C(═O)N(R^(B4))—, L^(B1) is directly        attached to the carbon atom labeled with 4;

L^(B2) is —O—, —S—, —NR^(B5)—, or —C(R^(B6))₂—, wherein R^(B5) ishydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; and each instance of R^(B6) is independently hydrogen,halogen, or substituted or unsubstituted alkyl;

each of X^(B1), X^(B2), and X^(B3) is independently N or CR^(B7),wherein each instance of R^(B7) is independently hydrogen, halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two R^(a)groups are joined to form a substituted or unsubstituted heterocyclic orsubstituted or unsubstituted heteroaryl ring;

each instance of R^(B1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

o1 is 0, 1, 2, 3, 4, or 5;

each instance of R^(B2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

o2 is 0, 1, 2, 3, or 4;

each instance of R^(B3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂, or an instance of R^(B3) and an instance of R^(B7)are joined to form a substituted or unsubstituted heterocyclic ring; and

o3 is 0, 1, or 2.

In certain embodiments, L^(B1) is —N(R^(B4))C(═O)— (e.g., —NHC(═O)—) or—C(═O)N(R^(B4))— (e.g., —C(═O)NH—). In certain embodiments, L^(B1) is—N(R^(B4))C(═O)N(R^(B4))—(e.g., —NHC(═O)NH)—).

In certain embodiments, all instances of R^(B4) are the same. In certainembodiments, two instances of R^(B4) are different from each other. Incertain embodiments, at least one instance of R^(B4) is hydrogen. Incertain embodiments, at least one instance of R^(B4) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(B4) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, L^(B2) is —O— or —S—. In certain embodiments,L^(B2) is —NR^(B5)— (e.g., —NH—). In certain embodiments, L^(B2) is—C(R^(B6))₂— (e.g., —CH₂—).

In certain embodiments, R^(B5) is hydrogen. In certain embodiments,R^(B5) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(B5) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(B6) are the same. In certainembodiments, two instances of R^(B6) are different from each other. Incertain embodiments, at least one instance of R^(B6) is hydrogen. Incertain embodiments, at least one instance of R^(B6) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(B6) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl).

In certain embodiments, X^(B1), X^(B2), and X^(B3) are CR^(B7) (e.g.,CH), N, and N, respectively. In certain embodiments, each of X^(B1),X^(B2), and X^(B3) is CR^(B7) (e.g., CH). In certain embodiments,X^(B1), X^(B2), and X^(B3) are N, CR^(B7) (e.g., CH), and CR^(B7) (e.g.,CH), respectively.

In certain embodiments, all instances of R^(B7) are the same. In certainembodiments, two instances of R^(B7) are different from each other. Incertain embodiments, at least one instance of R^(B7) is hydrogen. Incertain embodiments, at least one instance of R^(B7) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(B7) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(B7) is —CH₃. In certain embodiments, at least one instance ofR^(B7) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(B7) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(B7) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(B7) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(B7) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(B7) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(B7) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(B7) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(B7) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(B7) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(B7) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(B7)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(B7) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(B7) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(B7) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(B7) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, all instances of R^(B1) are the same. In certainembodiments, two instances of R^(B1) are different from each other. Incertain embodiments, at least one instance of R^(B1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(B1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(B1) is —CF₃ or —C(Me)₂CN. In certain embodiments, at least oneinstance of R^(B1) is —CH₃, unsubstituted ethyl, perfluoroethyl,unsubstituted propyl, perfluoropropyl, unsubstituted butyl, orperfluorobutyl. In certain embodiments, at least one instance of R^(B1)is substituted or unsubstituted alkenyl (e.g., substituted orunsubstituted C₁₋₆ alkenyl). In certain embodiments, at least oneinstance of R^(B1) is substituted or unsubstituted alkynyl (e.g.,substituted or unsubstituted C₁₋₆ alkynyl). In certain embodiments, atleast one instance of R^(B1) is substituted or unsubstituted carbocyclyl(e.g., substituted or unsubstituted, monocyclic, 3- to 7-memberedcarbocyclyl). In certain embodiments, at least one instance of R^(B1) issubstituted or unsubstituted heterocyclyl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heterocyclyl, wherein one,two, or three atoms in the heterocyclic ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, at least oneinstance of R^(B1) is substituted or unsubstituted aryl (e.g.,substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, at least one instance of R^(B1) is substituted orunsubstituted phenyl. In certain embodiments, at least one instance ofR^(B1) is substituted or unsubstituted heteroaryl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, at least oneinstance of R^(B1) is substituted or unsubstituted imidazolyl (e.g.,substituted or unsubstituted 1-imidazolyl). In certain embodiments, atleast one instance of R^(B1) is —OR^(a) (e.g., —OH, —O(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or—O(substituted or unsubstituted phenyl) (e.g., —OPh)). In certainembodiments, at least one instance of R^(B1) is —SR^(a) (e.g., —SH,—S(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —SMe, —SEt, —SPr,—SBu, or —SBn), or —S(substituted or unsubstituted phenyl) (e.g.,—SPh)). In certain embodiments, at least one instance of R^(B1) is—N(R^(a))₂ (e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NHMe), or —N(substituted or unsubstituted C₁₋₆alkyl)-(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). Incertain embodiments, at least one instance of R^(B1) is —CN, —SCN, or—NO₂. In certain embodiments, at least one instance of R^(B1) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, at least one instance of R^(B1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(B1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(B1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(B1) is halogen,substituted or unsubstituted alkyl, or substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur.

In certain embodiments, o1 is 0. In certain embodiments, o1 is 1. Incertain embodiments, o1 is 2. In certain embodiments, o1 is 3. Incertain embodiments, o1 is 4. In certain embodiments, o1 is 5.

In certain embodiments, all instances of R^(B2) are the same. In certainembodiments, two instances of R^(B2) are different from each other. Incertain embodiments, at least one instance of R^(B2) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(B2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(B2) is —CH₃. In certain embodiments, at least one instance ofR^(B2) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(B2) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(B2) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(B2) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(B2) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(B2) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(B2) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(B2) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(B2) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(B2) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(B2) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(B2)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(B2) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(B2) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(B2) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(B2) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, o2 is 0. In certain embodiments, o2 is 1. Incertain embodiments, o2 is 2. In certain embodiments, o2 is 3. Incertain embodiments, o2 is 4.

In certain embodiments, all instances of R^(B3) are the same. In certainembodiments, two instances of R^(B3) are different from each other. Incertain embodiments, at least one instance of R^(B3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(B3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(B3) is —CH₃. In certain embodiments, at least one instance ofR^(B3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(B3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(B3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(B3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(B3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(B3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(B3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(B3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(B3) issubstituted or unsubstituted pyridyl (e.g., 3-pyridyl). In certainembodiments, at least one instance of R^(B3) is —OR^(a) (e.g., —OH, O(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —OMe, —OEt, —OPr,—OBu, or —OBn), or —O(substituted or unsubstituted phenyl) (e.g.,—OPh)). In certain embodiments, at least one instance of R^(B3) is—SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted or unsubstitutedphenyl) (e.g., —SPh)). In certain embodiments, at least one instance ofR^(B3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆alkyl) (e.g., —NHMe), or —N(substituted or unsubstituted C₁₋₆alkyl)-(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). Incertain embodiments, at least one instance of R^(B3) is —CN, —SCN, or—NO₂. In certain embodiments, at least one instance of R^(B3) is—C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certainembodiments, at least one instance of R^(B3) is —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. In certain embodiments, atleast one instance of R^(B3) is —OC(═O)R^(a), —OC(═O)OR^(a), or—OC(═O)N(R^(a))₂. In certain embodiments, an instance of R^(B3) and aninstance of R^(B7) are joined to form a substituted or unsubstitutedheterocyclic ring (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, an instance of R^(B3) and an instance of R^(B7)are joined to form

In certain embodiments, at least one instance of R^(B3) is substitutedor unsubstituted heteroaryl or —C(═O)N(R^(a))₂, or an instance of R^(B3)and an instance of R^(B7) are joined to form a substituted orunsubstituted, monocyclic, 5- to 6-membed heterocyclic ring, whereinone, two, or three atoms in the heterocyclic ring system areindependently nitrogen, oxygen, or sulfur.

In certain embodiments, o3 is 0. In certain embodiments, o3 is 1. Incertain embodiments, o3 is 2.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein each of R^(B9) and R^(B10) is independentlysubstituted or unsubstituted, monocyclic, 5- to 6-membered heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur. In certain embodiments,R^(B9) is substituted or unsubstituted imidazolyl (e.g., substituted orunsubstituted 1-imidazolyl). In certain embodiments, R^(B10) issubstituted or unsubstituted pyridyl (e.g., substituted or unsubstituted3-pyridyl).

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein Ring B is a substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclic ring, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (B) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (B) is not AMN-107,AZ-628, BAY 73-4506, BAY-439006, or a pharmaceutically acceptable salt,solvate, hydrate, polymorph, co-crystal, tautomer, stereoisomer, orisotopically labeled derivative thereof.

It is believed that compounds of Formula (B), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are ANL inhibitors (e.g., AMN-107), BRAF inhibitors (e.g.,AZ-628 and BAY-439006), VEGFR inhibitors (e.g., VEGFR2 inhibitors, suchas BAY 73-4506), and/or TIE2 inhibitors (e.g., BAY 73-4506).

Compounds of Formula (C)

In another aspect, the present disclosure provides compounds of Formula(C):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

Ring C1 is a substituted or unsubstituted, monocyclic, 5- to 6-memberedheterocyclic ring or a substituted or unsubstituted, monocyclic, 5- to6-membered heteroaryl ring, wherein one or two atoms in the heterocyclicor heteroaryl ring system are nitrogen;

Ring C2 is a substituted or unsubstituted, monocyclic, 6-memberedcarbocyclic ring or a substituted or unsubstituted phenyl ring;

Ring C3 is a substituted or unsubstituted, monocyclic, 5- to 6-memberedheterocyclic ring or a substituted or unsubstituted, monocyclic, 5- to6-membered heteroaryl ring, wherein one or two atoms in the heterocyclicor heteroaryl ring system are nitrogen;

each instance of R^(C1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

p1 is 0, 1, 2, 3, 4, 5, 6, 7, 8, or 9;

each instance of R^(C2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂, or two instances ofR^(C2) are joined to form a substituted or unsubstituted carbocyclicring;

p2 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11;

each instance of R^(C3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

p3 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

In certain embodiments, Ring C1 is a substituted or unsubstituted,monocyclic, 5- to 6-membered heterocyclic ring, wherein one or two atomsin the heterocyclic ring system are nitrogen. In certain embodiments,Ring C1 is a substituted or unsubstituted 1,2-dihydropyridinyl ring. Incertain embodiments, Ring C1 is a substituted or unsubstituted,monocyclic, 5- to 6-membered heteroaryl ring, wherein one or two atomsin the heteroaryl ring system are nitrogen. In certain embodiments, RingC1 is a substituted or unsubstituted pyrazolyl ring, substituted orunsubstituted imidazolyl ring, substituted or unsubstituted pyridylring, or substituted or unsubstituted pyrimidinyl ring.

In certain embodiments, Ring C2 is a substituted or unsubstituted,monocyclic, 6-membered carbocyclic ring. In certain embodiments, Ring C2is a cyclohexa-2,4-dienone ring or cyclohexa-1,4-diene ring. In certainembodiments, Ring C2 is a substituted or unsubstituted phenyl ring.

In certain embodiments, Ring C3 is a substituted or unsubstituted,monocyclic, 5- to 6-membered heterocyclic ring, wherein one or two atomsin the heterocyclic ring system are nitrogen. In certain embodiments,Ring C3 is a substituted or unsubstituted piperidinyl ring (e.g.,substituted or unsubstituted 4-piperidinyl ring) or substituted orunsubstituted 1,4-dihydropyridinyl ring (e.g., substituted orunsubstituted 1,4-dihydropyridin-4-yl ring). In certain embodiments,Ring C3 is a substituted or unsubstituted, monocyclic, 5- to 6-memberedheteroaryl ring, wherein one or two atoms in the heteroaryl ring systemare nitrogen. In certain embodiments, Ring C3 is a substituted orunsubstituted pyridinyl ring (e.g., substituted or unsubstituted4-pyridinyl ring) or substituted or unsubstituted imidazolyl ring (e.g.,substituted or unsubstituted 2-imidazolyl ring).

In certain embodiments, all instances of R^(C1) are the same. In certainembodiments, two instances of R^(C1) are different from each other. Incertain embodiments, at least one instance of R^(C1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(C1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(C1) is —CH₂CH₂OH. In certain embodiments, at least one instance ofR^(C1) is —CH₃, —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(C1) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(C1) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(C1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(C1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(C1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(C1) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(C1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(C1) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(C1) is —OCH₂CH₂N(Me)₂. In certain embodiments, at least one instanceof R^(C1) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(C1) is —N(R^(a))₂ (e.g.,—NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe) or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(C1)is —NH₂. In certain embodiments, at least one instance of R^(C1) is —CN,—SCN, or —NO₂. In certain embodiments, at least one instance of R^(C1)is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. Incertain embodiments, at least one instance of R^(C1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(C1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(C1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(C1) is substitutedor unsubstituted alkyl, —OR^(a), —N(R^(a))₂, or —CN.

In certain embodiments, p1 is 0. In certain embodiments, p1 is 1. Incertain embodiments, p1 is 2. In certain embodiments, p1 is 3. Incertain embodiments, p1 is 4. In certain embodiments, p1 is 5. Incertain embodiments, p1 is 6. In certain embodiments, p1 is 7. Incertain embodiments, p1 is 8. In certain embodiments, p1 is 9.

In certain embodiments, all instances of R^(C2) are the same. In certainembodiments, two instances of R^(C2) are different from each other. Incertain embodiments, at least one instance of R^(C2) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(C2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(C2) is —CH₃. In certain embodiments, at least one instance ofR^(C2) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(C2) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(C2) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(C2) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(C2) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(C2) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(C2) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(C2) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(C2) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(C2) is —OCH₂(unsubstituted cyclopropyl). In certain embodiments, atleast one instance of R^(C2) is —SR^(a) (e.g., —SH, —S(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or—S(substituted or unsubstituted phenyl) (e.g., —SPh)). In certainembodiments, at least one instance of R^(C2) is —N(R^(a))₂ (e.g., —NH₂,—NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or—N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, atleast one instance of R^(C2) is —CN, —SCN, or —NO₂. In certainembodiments, at least one instance of R^(C2) is —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certain embodiments, atleast one instance of R^(C2) is —C(═O)R^(a), —C(═O)OR^(a), or—C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certainembodiments, at least one instance of R^(C2) is —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. In certain embodiments, atleast one instance of R^(C2) is —OC(═O)R^(a), —OC(═O)OR^(a), or—OC(═O)N(R^(a))₂. In certain embodiments, two instances of R^(C2) arejoined to form a substituted or unsubstituted carbocyclic ring (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed carbocyclicring). In certain embodiments, two instances of R^(C2) are joined toform a substituted or unsubstituted cyclopentyl ring or substituted orunsubstituted cyclopentenyl ring.

In certain embodiments, at least one instance of R^(C2) is halogen or—OR^(a).

In certain embodiments, p2 is 0. In certain embodiments, p2 is 1. Incertain embodiments, p2 is 2. In certain embodiments, p2 is 3. Incertain embodiments, p2 is 4. In certain embodiments, p2 is 5. Incertain embodiments, p2 is 6. In certain embodiments, p2 is 7. Incertain embodiments, p2 is 8. In certain embodiments, p2 is 9. Incertain embodiments, p2 is 10. In certain embodiments, p2 is 11.

In certain embodiments, all instances of R^(C3) are the same. In certainembodiments, two instances of R^(C3) are different from each other. Incertain embodiments, at least one instance of R^(C3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(C3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(C3) is —CH₃. In certain embodiments, at least one instance ofR^(C3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(C) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(C3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(C3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(C3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(C3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(C3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(C3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(C3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(C3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(C3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(C3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(C3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(C3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(C3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(C3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, p3 is 0. In certain embodiments, p3 is 1. Incertain embodiments, p3 is 2. In certain embodiments, p3 is 3. Incertain embodiments, p3 is 4. In certain embodiments, p3 is 5. Incertain embodiments, p3 is 6. In certain embodiments, p3 is 7. Incertain embodiments, p3 is 8. In certain embodiments, p3 is 9. Incertain embodiments, p3 is 10.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein Ring C4 is a substituted or unsubstituted,monocyclic, 5- to 6-membed carbocyclic ring (e.g., substituted orunsubstituted cyclopentyl ring or substituted or unsubstitutedcyclopentenyl ring).

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein Ring C5 is a substituted or unsubstituted,monocyclic, 5- to 6-membed carbocyclic ring (e.g., substituted orunsubstituted cyclopentenyl ring).

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (C) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (C) is not GDC-0879,KIN001-260, SB590885, CHIR99021, or a pharmaceutically acceptable salt,solvate, hydrate, polymorph, co-crystal, tautomer, stereoisomer, orisotopically labeled derivative thereof.

It is believed that compounds of Formula (C), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are BRAF inhibitors (e.g., GDC-0879 and SB590885), IKKinhibitors (e.g., IKKI3 inhibitors, such as KIN001-260), and/or GSK3inhibitors (e.g., GSK3B inhibitors, such as CHIR99021).

Compounds of Formula (D)

In another aspect, the present disclosure provides compounds of Formula(D):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(D1) is a substituted or unsubstituted, saturated or unsaturated, C₃₋₇aliphatic chain, optionally wherein one, two, or three chain atoms ofthe aliphatic chain are independently replaced with —O—, —S—, —NR^(D6)—,—N═, or ═N—, wherein each instance of R^(D6) is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;

each instance of R^(D1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

q1 is 0, 1, 2, 3, 4, or 5;

each instance of R^(D2) is independently hydrogen, halogen, orsubstituted or unsubstituted C₁₋₆ alkyl;

each instance of R^(D3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

q3 is 0, 1, 2, or 3;

each instance of R^(D4) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

q4 is 0, 1, 2, or 3; and

each instance of R^(D5) is independently hydrogen, substituted orunsubstituted C₁₋₆ alkyl, or a nitrogen protecting group.

In certain embodiments, L^(D1) is a substituted or unsubstituted,saturated or unsaturated, C₅ aliphatic chain, optionally wherein one,two, or three chain atoms of the aliphatic chain are independentlyreplaced with —O—, —S—, —NR^(D6)— (e.g., —NH—), —N═, or ═N—In certainembodiments, L^(D1) is of the formula:

wherein each instance of R^(D7) is independently hydrogen, halogen,substituted or unsubstituted C₁₋₆ alkyl, or substituted or unsubstitutedphenyl; and L^(D2) is —O—, —S—, —N(R^(D8))—, or —C(R^(D9))₂—, whereinR^(D8) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group, and each instance of R^(D9) is independentlyhydrogen, halogen, or substituted or unsubstituted C₁₋₆ alkyl. Incertain embodiments, L^(D1) is of the formula:

wherein R^(D7) is substituted or unsubstituted phenyl. In certainembodiments, L^(D1) is a substituted or unsubstituted, saturated orunsaturated, C₃, C₄, C₆, or C₇ aliphatic chain, optionally wherein one,two, or three chain atoms of the aliphatic chain are independentlyreplaced with —O—, —S—, —NR^(D6)— (e.g., —NH—), —N═, or ═N—.

In certain embodiments, all instances of R^(D6) are the same. In certainembodiments, two instances of R^(D6) are different from each other. Incertain embodiments, at least one instance of R^(D6) is hydrogen. Incertain embodiments, at least one instance of R^(D6) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(D6) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(D1) are the same. In certainembodiments, two instances of R^(D1) are different from each other. Incertain embodiments, at least one instance of R^(D1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(D1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(D1) is —CH₃. In certain embodiments, at least one instance ofR^(D1) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(D1) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(D1) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(D1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(D1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(D1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(D1) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(D1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(D1) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(D1) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(D1) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(D1)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(D1) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(D1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(D1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(D1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, q1 is 0. In certain embodiments, q1 is 1. Incertain embodiments, q1 is 2. In certain embodiments, q1 is 3. Incertain embodiments, q1 is 4. In certain embodiments, q1 is 5.

In certain embodiments, both instances of R^(D2) are the same. Incertain embodiments, two instances of R^(D2) are different from eachother. In certain embodiments, at least one instance of R^(D2) ishydrogen. In certain embodiments, each instance of R^(D2) is hydrogen.In certain embodiments, at least one instance of R^(D2) is halogen(e.g., F, Cl, Br, or I). In certain embodiments, at least one instanceof R^(D2) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl).

In certain embodiments, all instances of R^(D3) are the same. In certainembodiments, two instances of R^(D3) are different from each other. Incertain embodiments, at least one instance of R^(D3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(D3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(D3) is —CH₃. In certain embodiments, at least one instance ofR^(D3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(D3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(D3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(D3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(D3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(D3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(D3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(D3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(D3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(D3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(D3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —Nme₂)). In certain embodiments, at least one instance of R^(D3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(D3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(D3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)Nme₂). In certain embodiments, at least one instance of R^(D3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(D3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, q3 is 0. In certain embodiments, q3 is 1. Incertain embodiments, q3 is 2. In certain embodiments, q3 is 3.

In certain embodiments, all instances of R^(D4) are the same. In certainembodiments, two instances of R^(D4) are different from each other. Incertain embodiments, at least one instance of R^(D4) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(D4) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(D4) is —CH₃. In certain embodiments, at least one instance ofR^(D4) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(D4) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(D4) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(D4) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(D4) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(D4) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(D4) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(D4) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(D4) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(D4) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(D4) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —Nme₂)). In certain embodiments, at least one instance of R^(D4)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(D4) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(D4) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)Nme₂). In certain embodiments, at least one instance of R^(D4) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(D4) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, q4 is 0. In certain embodiments, q4 is 1. Incertain embodiments, q4 is 2. In certain embodiments, q4 is 3.

In certain embodiments, both instances of R^(D5) are the same. Incertain embodiments, two instances of R^(D5) are different from eachother. In certain embodiments, at least one instance of R^(D5) ishydrogen. In certain embodiments, each instance of R^(D5) is hydrogen.In certain embodiments, at least one instance of R^(D5) is substitutedor unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(D5) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, the compound of Formula (D) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein R^(D7) is substituted or unsubstituted phenyl.

In certain embodiments, the compound of Formula (D) is of the formula:

or a solvate, hydrate, polymorph, co-crystal, tautomer, stereoisomer,isotopically labeled derivative, or prodrug thereof.

In certain embodiments, a compound of Formula (D) is not KIN001-244, ora pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (D), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are PDK inhibitors.

Compounds of Formula (E)

In another aspect, the present disclosure provides compounds of Formula(E):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

X^(E1) is —O—, —S—, —C(═O)—, —C(═S)—, or —C(═NR^(E4))—, wherein R^(E4)is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group;

X^(E2) is —O—, —S—, —C(═O)—, or —C(═S)—;

R^(E1) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂, or R^(E1) and R^(E4) are joined to form asubstituted or unsubstituted heterocyclic ring or a substituted orunsubstituted heteroaryl ring;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

each instance of R^(E2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

r2 is 0, 1, 2, or 3;

each instance of R^(E3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

r3 is 0, 1, 2, 3, or 4.

In certain embodiments, X^(E1) is —S—. In certain embodiments, X^(E1) is—C(═NR⁴)—. In certain embodiments, X^(E1) is —O—, —C(═O)—, or —C(═S)—.

In certain embodiments, R^(E4) is hydrogen. In certain embodiments,R^(E) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, RM is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, X^(E2) is —S—. In certain embodiments, X^(E2) is—C(═O)—. In certain embodiments, X^(E2) is —O— or —C(═S)—.

In certain embodiments, R^(E1) is hydrogen. In certain embodiments,R^(E1) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(E1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(E1) is —CH₃. Incertain embodiments, R^(E1) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(E1) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(E1) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(E1) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(E1) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(E1) is a substituted or unsubstituted4H-pyran-4-one moiety. In certain embodiments, R^(E1) is substituted orunsubstituted aryl (e.g., substituted or unsubstituted, 6- to10-membered aryl). In certain embodiments, R^(E1) is substituted orunsubstituted phenyl. In certain embodiments, R^(E1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, R^(E1) is —OR^(a) (e.g., —OH,—O(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —OMe, —OEt, —OPr,—OBu, or —OBn), or —O(substituted or unsubstituted phenyl) (e.g.,—OPh)). In certain embodiments, R^(E1) is —SR^(a) (e.g., —SH,—S(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —SMe, —SEt, —SPr,—SBu, or —SBn), or —S(substituted or unsubstituted phenyl) (e.g.,—SPh)). In certain embodiments, R^(E1) is —N(R^(a))₂ (e.g., —NH₂,—NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or—N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, R^(E1)is —CN, —SCN, or —NO₂. In certain embodiments, R^(E1) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, R^(E1) is —C(═O)R^(a), —C(═O)OR^(a), or —C(═O)N(R^(a))₂(e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments,R^(E1) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or—NR^(a)C(═O)N(R^(a))₂. In certain embodiments, R^(E1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂. In certain embodiments, R^(E1) andR^(E) are joined to form a substituted or unsubstituted heterocyclicring (e.g., substituted or unsubstituted, monocyclic, 5- to 6-membedheterocyclic ring, wherein one, two, or three atoms in the heterocyclicring system are independently nitrogen, oxygen, or sulfur) or asubstituted or unsubstituted heteroaryl ring (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl ring, wherein one,two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur).

In certain embodiments, all instances of R^(E2) are the same. In certainembodiments, two instances of R^(E2) are different from each other. Incertain embodiments, at least one instance of R^(E2) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(E2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(E2) is —CH₃. In certain embodiments, at least one instance ofR^(E2) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(E2) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(E2) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(E2) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(E2) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(E2) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(E2) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(E2) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(E2) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(E2) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(E2) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(E2)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(E2) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(E2) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(E2) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(E2) is —OC(═O)R^(a),OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, r2 is 0. In certain embodiments, r2 is 1. Incertain embodiments, r2 is 2. In certain embodiments, r2 is 3.

In certain embodiments, all instances of R^(E3) are the same. In certainembodiments, two instances of R^(E3) are different from each other. Incertain embodiments, at least one instance of R^(E3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(E3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(E3) is —CH₃. In certain embodiments, at least one instance ofR^(E3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(E3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(E3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(E3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(E3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(E3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(E3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(E3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(E3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(E3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(E3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(E3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(E3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(E3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(E3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(E3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, r3 is 0. In certain embodiments, r3 is 1. Incertain embodiments, r3 is 2. In certain embodiments, r3 is 3. Incertain embodiments, r3 is 4.

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein:

each instance of R^(E5) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

r5 is 0, 1, 2, or 3.

In certain embodiments, all instances of R^(E5) are the same. In certainembodiments, two instances of R^(E5) are different from each other. Incertain embodiments, at least one instance of R^(E5) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(E5) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(E5) is —CH₃. In certain embodiments, at least one instance ofR^(E5) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(E5) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(E5) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(E5) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(E5) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(E5) is substituted or unsubstituted morpholinyl (e.g., substituted orunsubstituted N-morpholinyl). In certain embodiments, at least oneinstance of R^(E5) is substituted or unsubstituted aryl (e.g.,substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, at least one instance of R^(E5) is substituted orunsubstituted phenyl. In certain embodiments, at least one instance ofR^(E5) is substituted or unsubstituted heteroaryl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, at least oneinstance of R^(E5) is —OR^(a)(e.g., —OH, —O(substituted or unsubstitutedC₁₋₆ alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substitutedor unsubstituted phenyl) (e.g., —OPh)). In certain embodiments, at leastone instance of R^(E5) is —SR^(a) (e.g., —SH, —S(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or—S(substituted or unsubstituted phenyl) (e.g., —SPh)).

In certain embodiments, at least one instance of R^(E5) is —N(R^(a))₂(e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe),or —N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, atleast one instance of R^(E5) is —CN, —SCN, or —NO₂. In certainembodiments, at least one instance of R^(E5) is —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certain embodiments, atleast one instance of R^(E5) is —C(═O)R^(a), —C(═O)OR^(a), or—C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certainembodiments, at least one instance of R^(E5) is —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. In certain embodiments, atleast one instance of R^(E5) is —OC(═O)R^(a), —OC(═O)OR^(a), or—OC(═O)N(R^(a))₂.

In certain embodiments, r5 is 0. In certain embodiments, r5 is 1. Incertain embodiments, r5 is 2. In certain embodiments, r5 is 3.

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein X^(E3) is C or N, and Ring E is a substitutedor unsubstituted, monocyclic, 5- to 6-membered heterocyclic ring or asubstituted or unsubstituted, monocyclic, 5- to 6-membered heteroarylring, wherein one, two, or three atoms in the heterocyclic or heteroarylring system are independently nitrogen, oxygen, or sulfur.

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein R^(E6) is hydrogen, substituted orunsubstituted C₁₋₆ alkyl (e.g., CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl), or a nitrogen protecting group (e.g., Bn,Boc, Cbz, Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, the compound of Formula (E) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (E) is not KU55933,SP600125, or a pharmaceutically acceptable salt, solvate, hydrate,polymorph, co-crystal, tautomer, stereoisomer, or isotopically labeledderivative thereof.

It is believed that compounds of Formula (E), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are ATM inhibitors (e.g., KU55933) and/or JNK inhibitors (e.g.,SP600125).

Compounds of Formula (F)

In another aspect, the present disclosure provides compounds of Formula(F):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

X^(F) is —N(R^(F9))—, ═N—, —C(R^(F10))₂—, or ═C(R^(F10))—, wherein:

R^(F9) is hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group; and

each instance of R^(F10) is independently hydrogen, halogen, substitutedor unsubstituted alkyl, substituted or unsubstituted alkenyl,substituted or unsubstituted alkynyl, substituted or unsubstitutedcarbocyclyl, substituted or unsubstituted heterocyclyl, substituted orunsubstituted aryl, substituted or unsubstituted heteroaryl, —OR^(a),—N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

R^(F1) is absent, hydrogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, or a nitrogen protecting group;

R^(F2) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂;

R^(F3) is absent, hydrogen, halogen, or substituted or unsubstitutedC₁₋₆ alkyl, or R^(F3) and R^(F2) are joined to form ═O;

R^(F4) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂;

R^(F5) is absent, hydrogen, halogen, or substituted or unsubstitutedC₁₋₆ alkyl;

R^(F6) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂;

R^(F7) is hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group; and

R^(F8) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group.

In certain embodiments, X^(F) is —N(R^(F9))— or ═N—. In certainembodiments, X^(F) is —C(R^(F10))₂— or ═C(R^(F10))—.

In certain embodiments, R^(F9) is hydrogen. In certain embodiments,R^(F9) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, R^(F9) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(F9) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(F9) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(F9) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(F9) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(F9) is substituted or unsubstituted phenyl (e.g.,

In certain embodiments, R^(F9) is substituted or unsubstitutedheteroaryl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heteroaryl, wherein one, two, three, or four atoms in theheteroaryl ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(F9) is a nitrogen protecting group (e.g., Bn,Boc, Cbz, Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, two instances of R^(F10) are the same. Incertain embodiments, two instances of R^(F10) are different from eachother. In certain embodiments, at least one instance of R^(F10) ishalogen (e.g., F, Cl, Br, or I). In certain embodiments, at least oneinstance of R^(F10) is substituted or unsubstituted alkyl (e.g.,substituted or unsubstituted C₁₋₆ alkyl). In certain embodiments, atleast one instance of R^(F10) is —CH₃. In certain embodiments, at leastone instance of R^(F10) is —CF₃, unsubstituted ethyl, perfluoroethyl,unsubstituted propyl, perfluoropropyl, unsubstituted butyl, orperfluorobutyl. In certain embodiments, at least one instance of R^(F10)is substituted or unsubstituted alkenyl (e.g., substituted orunsubstituted C₁₋₆ alkenyl). In certain embodiments, at least oneinstance of R^(F10) is substituted or unsubstituted alkynyl (e.g.,substituted or unsubstituted C₁₋₆ alkynyl). In certain embodiments, atleast one instance of R^(F10) is substituted or unsubstitutedcarbocyclyl (e.g., substituted or unsubstituted, monocyclic, 3- to7-membered carbocyclyl). In certain embodiments, at least one instanceof R^(F10) is substituted or unsubstituted heterocyclyl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclyl,wherein one, two, or three atoms in the heterocyclic ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments, atleast one instance of R^(F10) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, at least one instance of R^(F10) is substituted orunsubstituted phenyl

In certain embodiments, at least one instance of R^(F10) is substitutedor unsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(F10) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(F10) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R10 is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(F10)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(F10) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(F10) is —C(═O)R^(a)(e.g., —C(═O)Me), —C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂,—C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments, at least oneinstance of R^(F10) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or—NR^(a)C(═O)N(R^(a))₂. In certain embodiments, at least one instance ofR^(F10) is —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(F10) is substitutedor unsubstituted phenyl, or —C(═O)R^(a).

In certain embodiments, R^(F1) is absent. In certain embodiments, R^(F1)is hydrogen. In certain embodiments, R^(F1) is substituted orunsubstituted alkyl, such as substituted or unsubstituted C₁₋₆ alkyl(e.g., —CH₃, —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl). Incertain embodiments, R^(F1) is substituted or unsubstituted alkenyl(e.g., substituted or unsubstituted C₁₋₆ alkenyl). In certainembodiments, R^(F1) is substituted or unsubstituted alkynyl (e.g.,substituted or unsubstituted C₁₋₆ alkynyl). In certain embodiments,R^(F1) is substituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, R^(F1) is substituted or unsubstituted cyclopentyl. Incertain embodiments, R^(F1) is substituted or unsubstituted heterocyclyl(e.g., substituted or unsubstituted, monocyclic, 5- to 6-membedheterocyclyl, wherein one, two, or three atoms in the heterocyclic ringsystem are independently nitrogen, oxygen, or sulfur). In certainembodiments, R^(F1) is substituted or unsubstituted aryl (e.g.,substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(F1) is substituted or unsubstituted phenyl. In certainembodiments, R^(F1) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(F1) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts). In certainembodiments, R^(F1) is absent, substituted or unsubstituted alky, orsubstituted or unsubstituted, monocyclic, 5- to 6-membed carbocyclyl.

In certain embodiments, R^(F2) is hydrogen. In certain embodiments,R^(F2) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(F2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(F2) is —CH₃. Incertain embodiments, R^(F2) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(F2) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(F2) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(F2) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(F2) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(F2) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(F2) is substituted or unsubstituted phenyl. In certainembodiments, R^(F2) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(F2) is —OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted orunsubstituted phenyl) (e.g., —OPh)). In certain embodiments, R^(F2) is—SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted or unsubstitutedphenyl) (e.g., —SPh)). In certain embodiments, R^(F2) is —N(R^(a))₂(e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe),or —N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, R^(F2)is —CN, —SCN, or —NO₂. In certain embodiments, R^(F2) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, R^(F2) is —C(═O)R^(a), —C(═O)OR^(a), or —C(═O)N(R^(a))₂(e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments,R^(F2) is —NR^(a)C(═O)R^(a) or —NR^(a)C(═O)OR^(a). In certainembodiments, R^(F2) is —NR^(a)C(═O)N(R^(a))₂ (e.g.,—NHC(═O)NH(substituted or unsubstituted C₁₋₆ alkyl), such as—NHC(═O)NH(t-Bu)). In certain embodiments, R^(F2) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, R^(F3) is absent. In certain embodiments, R^(F3)is hydrogen. In certain embodiments, R^(F3) is halogen (e.g., F, Cl, Br,or I). In certain embodiments, R^(F3) is substituted or unsubstitutedalkyl (e.g., substituted or unsubstituted C₁₋₆ alkyl). In certainembodiments, R^(F3) is —CH₃. In certain embodiments, R^(F3) is —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl. In certainembodiments, R^(F3) and R^(F2) are joined to form ═O.

In certain embodiments, R^(F4) is hydrogen. In certain embodiments,R^(F4) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(F4) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(F4) is —CH₃. Incertain embodiments, R^(F4) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(F4) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(F4) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(F4) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(F4) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(F4) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(F4) is substituted or unsubstituted phenyl. In certainembodiments, R^(F4) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(F4) is —OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted orunsubstituted phenyl) (e.g., —OPh)). In certain embodiments, R^(F4) is—SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted or unsubstitutedphenyl) (e.g., —SPh)). In certain embodiments, R^(F4) is —N(R^(a))₂(e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe),or —N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, R^(F4)is —CN, —SCN, or —NO₂. In certain embodiments, R^(F4) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, R^(F4) is —C(═O)R^(a), —C(═O)OR^(a), or —C(═O)N(R^(a))₂(e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments,R^(F4) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or—NR^(a)C(═O)N(R^(a))₂. In certain embodiments, R^(F4) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂. In certain embodiments, R^(F4) ishydrogen or substituted or unsubstituted alkyl.

In certain embodiments, R^(F5) is absent. In certain embodiments, R^(F5)is hydrogen. In certain embodiments, R^(F5) is halogen (e.g., F, Cl, Br,or I). In certain embodiments, R^(F5) is substituted or unsubstitutedalkyl (e.g., substituted or unsubstituted C₁₋₆ alkyl). In certainembodiments, R^(F5) is —CH₃. In certain embodiments, R^(F5) is —CF5,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl.

In certain embodiments, R^(F6) is hydrogen. In certain embodiments,R^(F6) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(F6) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(F6) is —CH₃. Incertain embodiments, R^(F6) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(F6) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(F6) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(F6) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(F6) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(F6) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(F6) is substituted or unsubstituted phenyl. In certainembodiments, R^(F6) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(F6) is —OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted orunsubstituted phenyl) (e.g., —OPh)). In certain embodiments, R^(F6) is—SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted or unsubstitutedphenyl) (e.g., —SPh)). In certain embodiments, R^(F6) is —N(R^(a))₂(e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe),or —N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, R^(F6)is —CN, —SCN, or —NO₂. In certain embodiments, R^(F6) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, R^(F6) is —C(═O)R^(a), —C(═O)OR^(a), or —C(═O)N(R^(a))₂(e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments,R^(F6) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or—NR^(a)C(═O)N(R^(a))₂. In certain embodiments, R^(F6) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, R^(F7) is hydrogen. In certain embodiments,R^(F7) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, R^(F7) is of theformula:

In certain embodiments, R^(F7) is substituted or unsubstituted alkenyl(e.g., substituted or unsubstituted C₁₋₆ alkenyl). In certainembodiments, R^(F7) is substituted or unsubstituted alkynyl (e.g.,substituted or unsubstituted C₁₋₆ alkynyl). In certain embodiments,R^(F7) is substituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, R^(F7) is substituted or unsubstituted heterocyclyl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclyl,wherein one, two, or three atoms in the heterocyclic ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(F7) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, R^(F7)is substituted or unsubstituted phenyl. In certain embodiments, R^(F7)is substituted or unsubstituted heteroaryl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, R^(F7) issubstituted or unsubstituted pyridyl (e.g., substituted or unsubstituted2-pyridyl). In certain embodiments, R^(F7) is a nitrogen protectinggroup (e.g., Bn, Boc, Cbz, Fmoc, trifluoroacetyl, triphenylmethyl,acetyl, or Ts). In certain embodiments, R^(F7) is substituted orunsubstituted alkyl or substituted or unsubstituted, monocyclic, 5- to6-membed heteroaryl, wherein one, two, three, or four atoms in theheteroaryl ring system are independently nitrogen, oxygen, or sulfur.

In certain embodiments, R^(F8) is hydrogen. In certain embodiments,R^(F8) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(F8) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, the compound of Formula (F) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (F) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (F) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (F) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (F) is not PD0332991,PD173074, WZ-7043, or a pharmaceutically acceptable salt, solvate,hydrate, polymorph, co-crystal, tautomer, stereoisomer, or isotopicallylabeled derivative thereof.

It is believed that compounds of Formula (F), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are CDK inhibitors (e.g., CDK4 inhibitors, such as PD0332991),FGFR inhibitors (e.g., FGFR1 inhibitors, such as PD173074), and/or CSF1Rinhibitors (e.g., WZ-7043).

Compounds of Formula (G)

In another aspect, the present disclosure provides compounds of Formula(G):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

Ring G is a substituted or unsubstituted phenyl ring or a substituted orunsubstituted, monocyclic, 5- to 6-membered heteroaryl ring, wherein oneor two atoms in the heteroaryl ring system are independently nitrogen,oxygen, or sulfur;

each instance of R^(G1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

t1 is 0, 1, 2, 3, 4, or 5;

L^(G) is a bond, —O—, —S—, —N(R^(G5))—, or —C(R^(G6))₂—, wherein R^(G5)is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; and each instance of R^(G6) is independently hydrogen,halogen, or substituted or unsubstituted C₁₋₆ alkyl;

R^(G2) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl, orsubstituted or unsubstituted heteroaryl;

R^(G3) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(G4) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), —OC(═O)N(R^(a))₂, —S(═O)₂R^(a),—S(═O)₂OR^(a), or —S(═O)₂N(R^(a))₂; and

t4 is 0, 1, 2, 3, or 4.

In certain embodiments, Ring G is a substituted or unsubstituted phenylring.

In certain embodiments, Ring G is a substituted or unsubstituted,monocyclic, 5- to 6-membered heteroaryl ring, wherein one or two atomsin the heteroaryl ring system are independently nitrogen, oxygen, orsulfur. In certain embodiments, Ring G is substituted or unsubstitutedpyrrolyl ring.

In certain embodiments, all instances of R^(G1) are the same. In certainembodiments, two instances of R^(G1) are different from each other. Incertain embodiments, at least one instance of R^(G1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(G1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(G1) is —CH₃. In certain embodiments, at least one instance ofR^(G1) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(G1) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(G1) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(G1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(G1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(G1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(G1) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(G1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(G1) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(G1) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(G1) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(G1)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(G1) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(G1) is —C(═O)R^(a)or —C(═O)OR^(a). In certain embodiments, at least one instance of R^(G1)is —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, —C(═O)NMe₂,—C(═O)NHCH₂CH₂N(Et)₂, —C(═O)— (substituted or unsubstitutedN-pyrrolidinyl), or —C(═O)-(substituted or unsubstitutedN-piperazinyl)). In certain embodiments, at least one instance of R^(G1)is —NR^(a)C(═O)R^(a) (e.g., —N(substituted or unsubstituted C₁₋₆alkyl)-C(═O)(substituted or unsubstituted C₁₋₆ alkyl), such as

In certain embodiments, at least one instance of R^(G1) is—NR^(a)C(═O)OR^(a) or —NR^(a)C(═O)N(R^(a))₂. In certain embodiments, atleast one instance of R^(G1) is —OC(═O)R^(a), —OC(═O)OR^(a), or—OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(G1) is substitutedor unsubstituted alkyl, —C(═O)N(R^(a))₂, or —N(R^(a))C(═O)R^(a).

In certain embodiments, t1 is 0. In certain embodiments, t1 is 1. Incertain embodiments, t1 is 2. In certain embodiments, t1 is 3. Incertain embodiments, t1 is 4. In certain embodiments, t1 is 5.

In certain embodiments, L^(G) is a bond. In certain embodiments, L^(G)is —O—, —S—, or —C(R^(G6))₂— (e.g., —CH₂—). In certain embodiments,L^(G) is —N(R^(G5))— (e.g., —NH—).

In certain embodiments, R^(G5) is hydrogen. In certain embodiments,R^(G5) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(G5) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, both instances of R^(G6) are the same. Incertain embodiments, two instances of R^(G6) are different from eachother. In certain embodiments, at least one instance of R^(G6) ishydrogen. In certain embodiments, each instance of R^(G6) is hydrogen.In certain embodiments, at least one instance of R^(G6) is halogen(e.g., F, Cl, Br, or I). In certain embodiments, at least one instanceof R^(G6) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl).

In certain embodiments, R^(G2) is hydrogen. In certain embodiments,R^(G2) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(G2) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, R^(G2) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(G2) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R^(G2) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(G2) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(G2) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(G2) is substituted or unsubstituted phenyl. In certainembodiments, R^(G2) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur).

In certain embodiments, R^(G3) is hydrogen. In certain embodiments,R^(G3) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(G3) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(G4) are the same. In certainembodiments, two instances of R^(G4) are different from each other. Incertain embodiments, at least one instance of R^(G4) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(G4) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(G4) is —CH₃. In certain embodiments, at least one instance ofR^(G4) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(G4) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(G4) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(G4) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(G4) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(G4) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(G4) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(G4) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(G4) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(G4) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(G4) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(G4)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(G4) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(G4) is —C(═O)R^(a),—C(═O)OR^(a) (e.g., —C(═O)O(substituted or unsubstituted C₁₋₆ alkyl),such as —C(═O)OMe), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(G4) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(G4) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂. In certain embodiments, at least oneinstance of R^(G4) is —S(═O)₂R^(a) (e.g., —S(═O)₂ (substituted orunsubstituted C₁₋₆ alkyl), such as

—S(═O)₂OR^(a), or —S(═O)₂N(R^(a))₂ (e.g., —S(═O)₂N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted phenyl), such as

In certain embodiments, at least one instance of R^(G4) is halogen,—C(═O)OR^(a), —S(═O)₂R^(a), or —S(═O)₂N(R^(a))₂.

In certain embodiments, t4 is 0. In certain embodiments, t4 is 1. Incertain embodiments, t4 is 2. In certain embodiments, t4 is 3. Incertain embodiments, t4 is 4.

In certain embodiments, the compound of Formula (G) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (G) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (G) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (G) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (G) is not BIBF-1120,PHA-665752, SU11248, SU11274, or a pharmaceutically acceptable salt,solvate, hydrate, polymorph, co-crystal, tautomer, stereoisomer, orisotopically labeled derivative thereof.

It is believed that compounds of Formula (G), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are VEGFR inhibitors (e.g., SU11248 (a VEGFR1 inhibitor) andBIBF-1120), PDGFR inhibitors (e.g., BIBF-1120 and SU11248), FGFRinhibitors (e.g., BIBF-1120), MET inhibitors (e.g., PHA-665752 andSU11274), KIT inhibitors (e.g., SU11248), and/or FLT inhibitors (e.g.,FLT3 inhibitors, such as SU11248).

Compounds of Formula (H)

In another aspect, the present disclosure provides compounds of Formula(H):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(H) is a substituted or unsubstituted, saturated or unsaturated, C₁₋₄aliphatic chain, optionally wherein one or two chain atoms of thealiphatic chain are independently replaced with —O—, —S—, —NR^(H5)—,—N═, or ═N—, wherein each instance of R^(H5) is independently hydrogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group;

each instance of R^(H1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

u1 is 0, 1, 2, 3, 4, or 5;

R^(H2) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(H3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

u3 is 0, 1, or 2;

each instance of R^(H4) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

u4 is 0, 1, 2, 3, 4, or 5.

In certain embodiments, L^(H) is a substituted or unsubstituted C₁aliphatic chain. In certain embodiments, L^(H) is —O— or —S—. In certainembodiments, L^(H) is —NR^(H5)— (e.g., —NH—). In certain embodiments,L^(H) is a substituted or unsubstituted, saturated or unsaturated, C₃aliphatic chain, optionally wherein one or two chain atoms of thealiphatic chain are independently replaced with —O—, —S—,—NR^(H5)—(e.g., —NH—), ═N—, or ═N—. In certain embodiments, L^(H) is—NR^(H5)C(═O)O— (e.g., —NHC(═O)O)— or —N(substituted or unsubstitutedphenyl)C(═O)O)—) or —OC(═O)NR^(H5)— (e.g., —OC(═O)NH— or—OC(═O)N(substituted or unsubstituted phenyl)-). In certain embodiments,L^(H) is —NR^(H5)C(═O)NR^(H5—) (e.g., —NHC(═O)NH)—. In certainembodiments, L^(H) is a substituted or unsubstituted, saturated orunsaturated, C₂ or C₄ aliphatic chain, optionally wherein one or twochain atoms of the aliphatic chain are independently replaced with —O—,—S—, —NR^(H5)— (e.g., —NH—), —N═, or ═N—.

In certain embodiments, all instances of R^(H5) are the same. In certainembodiments, two instances of R^(H5) are different from each other. Incertain embodiments, at least one instance of R^(H5) is hydrogen. Incertain embodiments, at least one instance of R^(H5) is substituted orunsubstituted alkyl, such as substituted or unsubstituted C₁₋₆ alkyl(e.g., —CH₃, —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl). Incertain embodiments, at least one instance of R^(H5) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(H5) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(H5) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(H5) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(H5) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(H5) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(H5) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(H5) is anitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc, trifluoroacetyl,triphenylmethyl, acetyl, or Ts). In certain embodiments, at least oneinstance of R^(H5) is hydrogen or substituted or unsubstituted phenyl.

In certain embodiments, all instances of R^(H1) are the same. In certainembodiments, two instances of R^(H1) are different from each other. Incertain embodiments, at least one instance of R^(H1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(H1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(H1) is —CH₃. In certain embodiments, at least one instance ofR^(H1) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(H1) is of the formula:

In certain embodiments, at least one instance of R^(H1) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, at least one instance of R^(H1) issubstituted or unsubstituted alkynyl (e.g., substituted or unsubstitutedC₁₋₆ alkynyl). In certain embodiments, at least one instance of R^(H1)is substituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(H1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(H1) is substituted or unsubstituted piperazinyl (e.g., substituted orunsubstituted N-piperazinyl, such as

In certain embodiments, at least one instance of R^(H1) is substitutedor unsubstituted aryl (e.g., substituted or unsubstituted, 6- to10-membered aryl). In certain embodiments, at least one instance ofR^(H1) is substituted or unsubstituted phenyl. In certain embodiments,at least one instance of R^(H1) is substituted or unsubstitutedheteroaryl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heteroaryl, wherein one, two, three, or four atoms in theheteroaryl ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, at least one instance of R^(H1) is —OR^(a)(e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —OMe,—OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstituted phenyl)(e.g., —OPh)). In certain embodiments, at least one instance of R^(H1)is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(H1) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(H1)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(H1) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(H1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(H1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(H1) is —OC(═O)R^(a),OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(H1) is substitutedor unsubstituted alkyl or substituted or unsubstituted, monocyclic, 5-to 6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur.

In certain embodiments, u1 is 0. In certain embodiments, u1 is 1. Incertain embodiments, u1 is 2. In certain embodiments, u1 is 3. Incertain embodiments, u1 is 4. In certain embodiments, u1 is 5.

In certain embodiments, R^(H2) is hydrogen. In certain embodiments,R^(H2) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(H2) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(H3) are the same. In certainembodiments, two instances of R^(H3) are different from each other. Incertain embodiments, at least one instance of R^(H3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(H3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(H3) is —CH₃. In certain embodiments, at least one instance ofR^(H3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(H3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(H3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(H3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(H3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(H3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(H3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(H3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(H3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(H3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(H3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(H3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(H3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(H3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(H3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(H3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, u3 is 0. In certain embodiments, u3 is 1. Incertain embodiments, u3 is 2. In certain embodiments, u3 is 3.

In certain embodiments, all instances of R^(H4) are the same. In certainembodiments, two instances of R^(H4) are different from each other. Incertain embodiments, at least one instance of R^(H4) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(H4) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(H4) is —CH₃. In certain embodiments, at least one instance ofR^(H4) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(H4) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(H4) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(H4) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(H4) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(H4) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(H4) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(H4) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(H4) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(H4) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(H4) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(H4)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(H4) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(H4) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe,—C(═O)NH(substituted or unsubstituted phenyl) (e.g.,—C(═O)NH-(2-chlorophenyl)), or —C(═O)NMe₂). In certain embodiments, atleast one instance of R^(H4) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),or —NR^(a)C(═O)N(R^(a))₂. In certain embodiments, at least one instanceof R^(H4) is —OC(═O)R^(a), OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, at least one instance of R^(H4) is substitutedor unsubstituted alkyl, or —C(═O)N(R^(a))₂.

In certain embodiments, u4 is 0. In certain embodiments, u4 is 1. Incertain embodiments, u4 is 2. In certain embodiments, u4 is 3. Incertain embodiments, u4 is 4. In certain embodiments, u4 is 5.

In certain embodiments, the compound of Formula (H) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (H) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (H) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (H) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (H) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

Compounds WH-4-023 and WH-4-025 are disclosed in U.S. Patent ApplicationPublication, US 2005/0026914, published Feb. 3, 2005, which isincorporated herein by reference. In certain embodiments, a compound ofFormula (H) is not KIN001-220, WH-4-023, or a pharmaceuticallyacceptable salt, solvate, hydrate, polymorph, co-crystal, tautomer,stereoisomer, or isotopically labeled derivative thereof. In certainembodiments, a compound of Formula (H) is not WH-4-025, or apharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (H), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are aurora kinase inhibitors (e.g., KIN001-220) and/or SRCinhibitors (e.g., WH-4-023).

Compounds of Formula (I)

In another aspect, the present disclosure provides compounds of Formula(I):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

R^(I1) is substituted or unsubstituted alkyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, or substituted or unsubstitutedheteroaryl;

R^(I2) is hydrogen or substituted or unsubstituted alkyl;

R^(I5) is hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted aralkyl, or substituted or unsubstituted carbocyclyl;

R^(I6) is hydrogen or substituted or unsubstituted alkyl;

each instance of R^(I7) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted aralkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —O-(substituted orunsubstituted alkyl), —NH-(substituted or unsubstituted alkyl),—NH-(substituted or unsubstituted aryl), —N(substituted or unsubstitutedalkyl)-(substituted or unsubstituted alkyl), —N(substituted orunsubstituted alkyl)-(substituted or unsubstituted aryl), —NO₂, or —CN;and

v7 is 0, 1, 2, 3, or 4.

In certain embodiments, R^(I1) is substituted or unsubstituted alkyl,such as substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(I1) is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(I1) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(I1) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(I1) is substituted or unsubstituted phenyl. In certainembodiments, R^(I1) is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur).

In certain embodiments, R^(I2) is hydrogen. In certain embodiments,R^(I2) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl).

In certain embodiments, R^(I5) is hydrogen. In certain embodiments,R^(I5) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(I5) is —CH₃. Incertain embodiments, R^(I5) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(I5) is substitutedor unsubstituted aralkyl (e.g., Bn). In certain embodiments, R^(I5) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed carbocyclyl).

In certain embodiments, R^(I6) is hydrogen. In certain embodiments,R^(I6) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl).

In certain embodiments, all instances of R^(I7) are the same. In certainembodiments, two instances of R^(I7) are different from each other. Incertain embodiments, at least one instance of R^(I7) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(I7) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(I7) is —CH₃. In certain embodiments, at least one instance ofR^(I7) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(I7) is substituted orunsubstituted aralkyl (e.g., Bn). In certain embodiments, at least oneinstance of R^(I7) is substituted or unsubstituted alkenyl (e.g.,substituted or unsubstituted C₁₋₆ alkenyl). In certain embodiments, atleast one instance of R^(I7) is substituted or unsubstituted carbocyclyl(e.g., substituted or unsubstituted, monocyclic, 3- to 7-memberedcarbocyclyl). In certain embodiments, at least one instance of R^(I7) issubstituted or unsubstituted heterocyclyl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heterocyclyl, wherein one,two, or three atoms in the heterocyclic ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, at least oneinstance of R^(I7) is substituted or unsubstituted aryl (e.g.,substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, at least one instance of R^(I7) is substituted orunsubstituted phenyl. In certain embodiments, at least one instance ofR^(I7) is substituted or unsubstituted heteroaryl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur). In certain embodiments, at least oneinstance of R^(I7) is —O-(substituted or unsubstituted alkyl) (e.g.,—OMe, —OEt, —OPr, or —OBu). In certain embodiments, at least oneinstance of R^(I7) is —NH-(substituted or unsubstituted alkyl) (e.g.,—NHMe), —NH-(substituted or unsubstituted aryl) (e.g., —NHPh),—N(substituted or unsubstituted alkyl)-(substituted or unsubstitutedalkyl) (e.g., —NMe₂), or —N(substituted or unsubstitutedalkyl)-(substituted or unsubstituted aryl). In certain embodiments, atleast one instance of R^(I7) is —NO₂ or —CN.

In certain embodiments, v7 is 0. In certain embodiments, v7 is 1. Incertain embodiments, v7 is 2. In certain embodiments, v7 is 3. Incertain embodiments, v7 is 4.

In certain embodiments, the compound of Formula (I) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof, wherein:

each instance of R^(I8) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

v8 is 0, 1, 2, 3, 4, or 5.

In certain embodiments, all instances of R^(I8) are the same. In certainembodiments, two instances of R^(I8) are different from each other. Incertain embodiments, at least one instance of R^(I8) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(I8) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(I8) is —CH₃. In certain embodiments, at least one instance ofR^(I8) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(I8) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(I8) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(I8) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(I8) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur).

In certain embodiments, at least one instance of R is substituted orunsubstituted piperidinyl (e.g., substituted or unsubstitutedN-piperidinyl, such as

or substituted or unsubstituted piperazinyl (e.g., substituted orunsubstituted N-piperazinyl, such as

In certain embodiments, at least one instance of R^(I8) is substitutedor unsubstituted aryl (e.g., substituted or unsubstituted, 6- to10-membered aryl). In certain embodiments, at least one instance ofR^(I8) is substituted or unsubstituted phenyl. In certain embodiments,at least one instance of R^(I8) is substituted or unsubstitutedheteroaryl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heteroaryl, wherein one, two, three, or four atoms in theheteroaryl ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, at least one instance of R^(I8) is —OR^(a)(e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —OMe,—OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstituted phenyl)(e.g., —OPh)). In certain embodiments, at least one instance of R^(I8)is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(I8) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(I8)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(I8) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(I8) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe,—C(═O)NMe₂, or —C(═O)-(substituted or unsubstituted N-piperidinyl)(e.g.,

certain embodiments, at least one instance of R^(I8) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(I8) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, v8 is 0. In certain embodiments, v8 is 1. Incertain embodiments, v8 is 2. In certain embodiments, v8 is 3. Incertain embodiments, v8 is 4. In certain embodiments, v8 is 5.

In certain embodiments, the compound of Formula (I) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (I) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (I) is not XMD 8-92, XMD11-50, XMD 8-85, or a pharmaceutically acceptable salt, solvate,hydrate, polymorph, co-crystal, tautomer, stereoisomer, or isotopicallylabeled derivative thereof.

It is believed that compounds of Formula (I), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are ERK inhibitors (e.g., ER^(K5) inhibitors, such as XMD 8-92,XMD11-50, and XMD 8-85) and/or LRRK inhibitors (e.g., LRR^(K2)inhibitors, such as XMD11-50).

Compounds of Formula (J)

In another aspect, the present disclosure provides compounds of Formula(J):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(J) is a substituted or unsubstituted, saturated or unsaturated, C₁₋₆aliphatic chain, optionally wherein one or two chain atoms of thealiphatic chain are independently replaced with —O—, —S—, —NR^(J6)—,—N═, or ═N—, wherein each instance of R^(J6) is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;

each instance of R^(J1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

w1 is 0, 1, 2, 3, or 4;

R^(J2) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

R^(J3) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂;

R^(J4) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(J5) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

w5 is 0, 1, 2, 3, 4, or 5.

In certain embodiments, L^(J) is a substituted or unsubstituted,saturated or unsaturated, C₃ aliphatic chain, optionally wherein one ortwo chain atoms of the aliphatic chain are independently replaced with—O—, —S—, —NR^(J6)— (e.g., —NH—), —N═, or ═N—. In certain embodiments,L^(J) is of the formula:

wherein each instance of R^(J7) is independently hydrogen, halogen(e.g., F, Cl, Br, or I), or substituted or unsubstituted C₁₋₆ alkyl(e.g., —CH₃, —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl). Incertain embodiments, all instances of R^(J7) are the same. In certainembodiments, two instances of R^(J7) are different from each other. Incertain embodiments, each instance of R^(J7) is hydrogen.

In certain embodiments, all instances of R^(J6) are the same. In certainembodiments, two instances of R^(J6) are different from each other. Incertain embodiments, at least one instance of R^(J6) is hydrogen. Incertain embodiments, at least one instance of R^(J6) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(J6) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(J1) are the same. In certainembodiments, two instances of R^(J1) are different from each other. Incertain embodiments, at least one instance of R^(J1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(J1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(J1) is —CH₃. In certain embodiments, at least one instance ofR^(J1) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(J1) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(J1) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(J1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(J1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(J1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(J1) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(J1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(J1) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(J1) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(J1) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(J1)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(J1) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(J1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(J1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(J1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, w1 is 0. In certain embodiments, w1 is 1. Incertain embodiments, w1 is 2. In certain embodiments, w1 is 3. Incertain embodiments, w1 is 4.

In certain embodiments, R^(J2) is hydrogen. In certain embodiments,R^(J2) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(J2) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, R^(J3) is hydrogen. In certain embodiments,R^(J3) is halogen (e.g., F, Cl, Br, or I). In certain embodiments,R^(J3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, R^(J3) is —CH₃. Incertain embodiments, R^(J3) is —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl. In certain embodiments, R^(J3) is substitutedor unsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆alkenyl). In certain embodiments, R^(J3) is substituted or unsubstitutedalkynyl (e.g., substituted or unsubstituted C₁₋₆ alkynyl). In certainembodiments, R is substituted or unsubstituted carbocyclyl (e.g.,substituted or unsubstituted, monocyclic, 3- to 7-membered carbocyclyl).In certain embodiments, R^(J3) is substituted or unsubstitutedheterocyclyl (e.g., substituted or unsubstituted, monocyclic, 5- to6-membed heterocyclyl, wherein one, two, or three atoms in theheterocyclic ring system are independently nitrogen, oxygen, or sulfur).In certain embodiments, R^(J3) is substituted or unsubstituted aryl(e.g., substituted or unsubstituted, 6- to 10-membered aryl). In certainembodiments, R^(J3) is substituted or unsubstituted phenyl. In certainembodiments, R is substituted or unsubstituted heteroaryl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heteroaryl,wherein one, two, three, or four atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(J3) is —OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆alkyl) (e.g., —OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted orunsubstituted phenyl) (e.g., —OPh)). In certain embodiments, R^(J3) is—SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted or unsubstitutedphenyl) (e.g., —SPh)). In certain embodiments, R^(J3) is —N(R^(a))₂(e.g., —NH₂, —NH(substituted or unsubstituted C₁₋₆ alkyl) (e.g., —NHMe),or —N(substituted or unsubstituted C₁₋₆ alkyl)-(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NMe₂)). In certain embodiments, R^(J3)is —CN, —SCN, or —NO₂. In certain embodiments, R^(J3) is—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂. In certainembodiments, R^(J3) is —C(═O)R^(a), —C(═O)OR^(a), or —C(═O)N(R^(a))₂(e.g., —C(═O)NH₂, —C(═O)NHMe, or —C(═O)NMe₂). In certain embodiments,R^(J3) is —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or—NR^(a)C(═O)N(R^(a))₂. In certain embodiments, R^(J3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, R^(J4) is hydrogen. In certain embodiments,R^(J4) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(J4) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(J5) are the same. In certainembodiments, two instances of R^(J5) are different from each other. Incertain embodiments, at least one instance of R^(J5) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(J5) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(J5) is —CH₃. In certain embodiments, at least one instance ofR^(J5) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(J5) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(J5) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(J5) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(J5) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(J5) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(J5) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(J5) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(J5) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(J5) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(J5) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(J5)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(J5) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(J5) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(J5) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(J5) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, w5 is 0. In certain embodiments, w5 is 1. Incertain embodiments, w5 is 2. In certain embodiments, w5 is 3. Incertain embodiments, w5 is 4. In certain embodiments, w5 is 5.

In certain embodiments, the compound of Formula (J) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (J) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (J) is not IM12, or apharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (J), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are GSK inhibitors (e.g., GSK3 inhibitors).

Compounds of Formula (K)

In another aspect, the present disclosure provides compounds of Formula(K):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

R^(K1) is hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, or substituted or unsubstitutedheteroaryl;

R^(K2) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(K3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

x3 is 0, 1, 2, 3, or 4;

R^(K4) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group;

each instance of R^(K5) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and

x5 is 0, 1, 2, 3, 4, or 5.

In certain embodiments, R^(K1) is hydrogen. In certain embodiments,R^(K1) is substituted or unsubstituted alkyl, such as substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, R^(K1) is C₁₋₆ alkylsubstituted with at least one —OH. In certain embodiments, R^(K1) is ofthe formula:

In certain embodiments, R is substituted or unsubstituted alkenyl (e.g.,substituted or unsubstituted C₁₋₆ alkenyl). In certain embodiments,R^(K1) is substituted or unsubstituted alkynyl (e.g., substituted orunsubstituted C₁₋₆ alkynyl). In certain embodiments, R^(K1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, R^(K1) is substituted or unsubstituted heterocyclyl (e.g.,substituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclyl,wherein one, two, or three atoms in the heterocyclic ring system areindependently nitrogen, oxygen, or sulfur). In certain embodiments,R^(K1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, R^(K1)is substituted or unsubstituted phenyl. In certain embodiments, R^(K1)is substituted or unsubstituted heteroaryl (e.g., substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur).

In certain embodiments, R^(K2) is hydrogen. In certain embodiments,R^(K2) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(K2) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(K3) are the same. In certainembodiments, two instances of R^(K3) are different from each other. Incertain embodiments, at least one instance of R^(K3) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(K3) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(K3) is —CH₃. In certain embodiments, at least one instance ofR^(K3) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(K3) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(K3) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(K3) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(K3) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(K3) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(K3) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(K3) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(K3) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(K3) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(K3) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(K3)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(K3) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(K3) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(K3) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(K3) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, x3 is 0. In certain embodiments, x3 is 1. Incertain embodiments, x3 is 2. In certain embodiments, x3 is 3. Incertain embodiments, x3 is 4.

In certain embodiments, R^(K4) is hydrogen. In certain embodiments,R^(K4) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, R^(K4) is a nitrogen protecting group (e.g., Bn, Boc, Cbz,Fmoc, trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(K5) are the same. In certainembodiments, two instances of R^(K5) are different from each other. Incertain embodiments, at least one instance of R^(K5) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(K5) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(K5) is —CH₃. In certain embodiments, at least one instance ofR^(K5) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(K5) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(K5) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(K5) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(K5) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(K5) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(K5) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(K5) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(K5) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(K5) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(K5) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(K5)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(K5) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(K5) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(K5) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(K5) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, x5 is 0. In certain embodiments, x5 is 1. Incertain embodiments, x5 is 2. In certain embodiments, x5 is 3. Incertain embodiments, x5 is 4. In certain embodiments, x5 is 5.

In certain embodiments, the compound of Formula (K) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (K) is not PD0325901, or apharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (K), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are MEK inhibitors.

Compounds of Formula (L)

In another aspect, the present disclosure provides compounds of Formula(L):

and pharmaceutically acceptable salts, solvates, hydrates, polymorphs,co-crystals, tautomers, stereoisomers, isotopically labeled derivatives,and prodrugs thereof, wherein:

L^(L) is a substituted or unsubstituted, saturated or unsaturated, C₁₋₄aliphatic chain, optionally wherein one or two chain atoms of thealiphatic chain are independently replaced with —O—, —S—, —NR^(L5)—,—N═, or ═N—, wherein each instance of R^(L5) is independently hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;

each instance of R^(L1) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring;

y1 is 0, 1, 2, 3, or 4;

each instance of R^(L2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;

y2 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10;

each instance of R^(L3) is independently hydrogen, halogen, orsubstituted or unsubstituted C₁₋₆ alkyl; and

each instance of R is independently hydrogen, substituted orunsubstituted C₁₋₆ alkyl, or a nitrogen protecting group.

In certain embodiments, L^(L) is a substituted or unsubstituted,saturated or unsaturated, C₂ aliphatic chain, optionally wherein onechain atom of the aliphatic chain is replaced with —O—, —S—, —NR^(L5)—(e.g., —NH—), —N═, or ═N—. In certain embodiments, L^(L) is—C(═O)NR^(L5)— (e.g., —C(═O)NH—) or —NR^(L5)C(═O)— (e.g., —NHC(═O)—). Incertain embodiments, L^(L) is a substituted or unsubstituted, saturatedor unsaturated, C₁, C₃, or C₄ aliphatic chain, optionally wherein one ortwo chain atoms of the aliphatic chain are independently replaced with—O—, —S—, —NR^(L5)— (e.g., —NH—), —N═, or ═N—.

In certain embodiments, all instances of R^(L5) are the same. In certainembodiments, two instances of R^(L5) are different from each other. Incertain embodiments, at least one instance of R^(L5) is hydrogen. Incertain embodiments, at least one instance of R^(L5) is substituted orunsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(L5) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, all instances of R^(L1) are the same. In certainembodiments, two instances of R^(L1) are different from each other. Incertain embodiments, at least one instance of R^(L1) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(L1) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(L1) is —CH₃. In certain embodiments, at least one instance ofR^(L1) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(L1) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(L1) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(L1) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(L1) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(L1) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(L1) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(L1) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(L1) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(L1) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(L1) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(L1)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(L1) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(L1) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(L1) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(L1) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, y1 is 0. In certain embodiments, y1 is 1. Incertain embodiments, y1 is 2. In certain embodiments, y1 is 3. Incertain embodiments, y1 is 4.

In certain embodiments, all instances of R^(L2) are the same. In certainembodiments, two instances of R^(L2) are different from each other. Incertain embodiments, at least one instance of R^(L2) is halogen (e.g.,F, Cl, Br, or I). In certain embodiments, at least one instance ofR^(L2) is substituted or unsubstituted alkyl (e.g., substituted orunsubstituted C₁₋₆ alkyl). In certain embodiments, at least one instanceof R^(L2) is —CH₃. In certain embodiments, at least one instance ofR^(L2) is —CF₃, unsubstituted ethyl, perfluoroethyl, unsubstitutedpropyl, perfluoropropyl, unsubstituted butyl, or perfluorobutyl. Incertain embodiments, at least one instance of R^(L2) is substituted orunsubstituted alkenyl (e.g., substituted or unsubstituted C₁₋₆ alkenyl).In certain embodiments, at least one instance of R^(L2) is substitutedor unsubstituted alkynyl (e.g., substituted or unsubstituted C₁₋₆alkynyl). In certain embodiments, at least one instance of R^(L2) issubstituted or unsubstituted carbocyclyl (e.g., substituted orunsubstituted, monocyclic, 3- to 7-membered carbocyclyl). In certainembodiments, at least one instance of R^(L2) is substituted orunsubstituted heterocyclyl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur). In certain embodiments, at least one instance ofR^(L2) is substituted or unsubstituted aryl (e.g., substituted orunsubstituted, 6- to 10-membered aryl). In certain embodiments, at leastone instance of R^(L2) is substituted or unsubstituted phenyl. Incertain embodiments, at least one instance of R^(L2) is substituted orunsubstituted heteroaryl (e.g., substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur). In certain embodiments, at least one instance of R^(L2) is—OR^(a) (e.g., —OH, —O(substituted or unsubstituted C₁₋₆ alkyl) (e.g.,—OMe, —OEt, —OPr, —OBu, or —OBn), or —O(substituted or unsubstitutedphenyl) (e.g., —OPh)). In certain embodiments, at least one instance ofR^(L2) is —SR^(a) (e.g., —SH, —S(substituted or unsubstituted C₁₋₆alkyl) (e.g., —SMe, —SEt, —SPr, —SBu, or —SBn), or —S(substituted orunsubstituted phenyl) (e.g., —SPh)). In certain embodiments, at leastone instance of R^(L2) is —N(R^(a))₂ (e.g., —NH₂, —NH(substituted orunsubstituted C₁₋₆ alkyl) (e.g., —NHMe), or —N(substituted orunsubstituted C₁₋₆ alkyl)-(substituted or unsubstituted C₁₋₆ alkyl)(e.g., —NMe₂)). In certain embodiments, at least one instance of R^(L2)is —CN, —SCN, or —NO₂. In certain embodiments, at least one instance ofR^(L2) is —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), or —C(═NR^(a))N(R^(a))₂.In certain embodiments, at least one instance of R^(L2) is —C(═O)R^(a),—C(═O)OR^(a), or —C(═O)N(R^(a))₂ (e.g., —C(═O)NH₂, —C(═O)NHMe, or—C(═O)NMe₂). In certain embodiments, at least one instance of R^(L2) is—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), or —NR^(a)C(═O)N(R^(a))₂. Incertain embodiments, at least one instance of R^(L2) is —OC(═O)R^(a),—OC(═O)OR^(a), or —OC(═O)N(R^(a))₂.

In certain embodiments, y2 is 0. In certain embodiments, y2 is 1. Incertain embodiments, y2 is 2. In certain embodiments, y2 is 3. Incertain embodiments, y2 is 4. In certain embodiments, y2 is 5. Incertain embodiments, y2 is 6. In certain embodiments, y2 is 7. Incertain embodiments, y2 is 8. In certain embodiments, y2 is 9. Incertain embodiments, y2 is 10.

In certain embodiments, both instances of R^(L3) are the same. Incertain embodiments, two instances of R^(L3) are different from eachother. In certain embodiments, at least one instance of R^(L3) ishydrogen. In certain embodiments, each instance of R^(L3) is hydrogen.In certain embodiments, at least one instance of R^(L3) is halogen(e.g., F, Cl, Br, or I). In certain embodiments, at least one instanceof R^(L3) is substituted or unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃,unsubstituted ethyl, perfluoroethyl, unsubstituted propyl,perfluoropropyl, unsubstituted butyl, or perfluorobutyl). In certainembodiments, each instance of R^(L3) is independently hydrogen orsubstituted or unsubstituted alkyl.

In certain embodiments, both instances of R^(L4) are the same. Incertain embodiments, two instances of R^(L4) are different from eachother. In certain embodiments, at least one instance of R^(L4) ishydrogen. In certain embodiments, each instance of R^(L4) is hydrogen.In certain embodiments, at least one instance of R^(L4) is substitutedor unsubstituted C₁₋₆ alkyl (e.g., —CH₃, —CF₃, unsubstituted ethyl,perfluoroethyl, unsubstituted propyl, perfluoropropyl, unsubstitutedbutyl, or perfluorobutyl). In certain embodiments, at least one instanceof R^(L4) is a nitrogen protecting group (e.g., Bn, Boc, Cbz, Fmoc,trifluoroacetyl, triphenylmethyl, acetyl, or Ts).

In certain embodiments, the compound of Formula (L) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (L) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, the compound of Formula (L) is of the formula:

or a pharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, isotopically labeled derivative, orprodrug thereof.

In certain embodiments, a compound of Formula (L) is not Y-27632, or apharmaceutically acceptable salt, solvate, hydrate, polymorph,co-crystal, tautomer, stereoisomer, or isotopically labeled derivativethereof.

It is believed that compounds of Formula (L), and pharmaceuticallyacceptable salts, solvates, hydrates, polymorphs, co-crystals,tautomers, stereoisomers, isotopically labeled derivatives, and prodrugsthereof, are ROCK inhibitors.

In another aspect, the present disclosure provides a mixture ofcompounds described herein. In certain embodiments, the mixture ofcompounds include two or more compounds of any one of Formula (A) to(L), and pharmaceutically acceptable salts, solvates, hydrates,polymorphs, co-crystals, tautomers, stereoisomers, isotopically labeledderivatives, and prodrugs thereof, wherein any two compounds of themixture of compounds are of the same or different Formula (e.g., any oneof Formula (A) to (L)).

Examples Materials and Methods Culture Conditions

Conventional (primed) human iPSC line C1 (Whitehead Institute Center forHuman Stem Cell Research, Cambridge, Mass.) (Hockemeyer et al., 2008)and human ESC lines WIBR2 and WIBR3 (Whitehead Institute Center forHuman Stem Cell Research, Cambridge, Mass.) (Lengner et al., 2010)(Lengner et al., 2010) were maintained on mitomycin C inactivated mouseembryonic fibroblast (MEF) feeder layers and passaged using Collagenase(1 mg/mL) or manual methods. Primed human ESCs and human iPSCs werecultured in human ESC medium [DMEM/F12 (Invitrogen) supplemented with15% fetal bovine serum (FBS) (Hyclone), 5% KnockOut Serum Replacement(Invitrogen), 1 mM glutamine (Invitrogen), 1% nonessential amino acids(Invitrogen), 0.1 mM b-mercaptoethanol (Sigma) and 4 ng/ml FGF2 (R&Dsystems)]. Naïve human ESCs/hiPSCs were kept on mitomycin C-inactivatedmouse embryonic fibroblast feeder cells, and were passaged by briefsingle-cell dissociation using Accutase (Gibco) every 5-10 days. Naïvehuman pluripotent cells were derived and maintained in serum-freeN2B27-based media. 500 mL of medium was generated by including: 240 mLDMEM/F12 (Invitrogen; 11320), 240 mL Neurobasal (Invitrogen; 21103), 5mL N2 supplement (Invitrogen; Ser. No. 17/502,048), 10 mL B27 supplement(Invitrogen; Ser. No. 17/504,044), 10 μg recombinant human LIF (madein-house), 1 mM glutamine (Invitrogen), 1% nonessential amino acids(Invitrogen), 0.1 mM 3-mercaptoethanol (Sigma), penicillin-streptomycin(Invitrogen), 5 mg/mL BSA (Sigma), and the following small molecules andcytokines: PD0325901 (Stemgent, 1 μM), IM-12 (Enzo, 1 μM), SB590885 (R&Dsystems, 0.5 μM), WH4-023 (A Chemtek, 1 μM), Y-27632 (Stemgent, 10 μM)and Activin A (Peprotech, 20 ng/mL). To accelerate the kinetics of naïvecell induction FGF2 (R&D systems, 8 ng/mL) and 0.5% KSR (Gibco) werealso included. Additional inhibitors described in this work include:CHIR99021 (Stemgent, 3 μM), SP600125 (R&D systems, 1 μM), PD173074(Stemgent, 0.1 μM), SB431542 (Tocris, 5 μM) and BIR^(B796) (AxonMedchem, 2 μM). Tissue culture media were filtered using a lowprotein-binding binding 0.22 μM filter (Corning). Alternativeformulations for naïve human ESC culture were followed as describedpreviously (Chan et al., 2013; Gafni et al., 2013; Valamehr et al.,2014; Ware et al., 2014).

Gene Targeting

Human ESCs and iPSCs were cultured in ROCK-inhibitor Y-27632 24 hoursprior to electroporation. Cell were harvested using 0.25% trypsin/EDTAsolution (Invitrogen) and 1×10⁷ cells resuspended in phosphate bufferedsaline (PBS) were electroporated if not otherwise indicated with 40 μgof donor plasmids and 5 g of each Talen encoding plasmid (Gene PulserXcell System, Bio-Rad: 250 V, 500 μF, 0.4 cm cuvettes (Costa et al.,2007)). Cells were subsequently plated on MEF feeder layers (DR4 MEFsfor puromycin selection) in human ESC medium supplemented withROCK-inhibitor for the first 24 hours. Individual colonies were pickedand expanded after puromycin selection (0.5 μg/ml) 10 to 14 days afterelectroporation. Correctly targeted clones were confirmed by southernblot (NdeI digested) and used for the removal of floxed PGK-purocassette. Cells were harvested using 0.25% trypsin/EDTA solution(Invitrogen) and 1×10⁷ cells resuspended in PBS were electroporated withpTurbo-Cre (40 jtg; GenBank accession number AF334827) (Gene PulserXcell System, Bio-Rad; 250 V, 500 jiF, 0.4-cm cuvettes). Cells weresubsequently plated on MEF feeder layers at a low density in human ESCmedium supplemented with ROCK inhibitor. Individual colonies were picked10-14 d after electroporation. The excision of PGK-puro was confirmed bySouthern blot analysis.

Lentiviral Infection

VSVG coated lentiviruses were generated in HEK-293 cells as describedpreviously (Brambrink et al., 2008; Soldner et al., 2009; Soldner etal., 2011). Briefly, culture medium was changed 12 hr post-transfectionand virus-containing supernatant was collected 48 to 72 hr posttransfection. Viral supernatant was filtered through a 0.45 mm filter.Virus-containing supernatants of the 2 reprogramming viruses(FUW-tetO-lox-hKLF2 and FUW-tetO-lox-hNANOG) were pooled andsupplemented with the FUW-lox-M2rtTA virus and an equal volume of freshculture medium. 1×10⁶ human ESCs were seeded 24 hr before transductionin T75 flasks on matrigel in mTesrl medium (STEMCELL Technologies). Twoconsecutive infections in the presence of 2 mg/ml of polybrene wereperformed over a period of 24 hr. Culture medium was changed 12 hr afterthe last infection. Five days after transduction, human ESCs werepassaged using 0.25% trypsin/EDTA solution (Invitrogen) and re-plated onmitomycin C inactivated mouse embryonic fibroblast (MEF) feeder layersin conventional human ESC medium. To induce conversion to the naïvestate, human ESCs were trypsinized and seeded at a density of 1×10⁵cells per 6 individual well on a MEF feeder layer in presence of ROCKinhibitor Y-27632. Medium was replaced 24 hr later with N2B27 basalmedium supplemented with PD0325901 (Stemgent, 1 μM), CHIR99021(Stemgent, 3 μM), 20 ng/mL hLIF (2i/L) and doxycycline (DOX)(Sigma-Aldrich; 2 mg/ml). OCT4-ΔPE-GFP+ human ESC colonies were pickedmanually within 10 days after DOX induction and passaged using Accutase(Gibco) on a MEF feeder layer. Upon the addition of 2i/L/DOX, latentOCT4-ΔPE-GFP-negative cells could be removed almost entirely byadditional treatment with 0.1 μM PD173074 and 5 μM SB431542, whicheffectively inhibit the signaling pathways on which primed human ESCsare reliant. These additional inhibitors also facilitated the isolationof transgene-dependent naïve human ESCs from wild-type WIBR3 human ESCs.

Chemical Screening

To screen for small molecules that support naïve human pluripotency,doxycycline was withdrawn from a clonal line of WIBR3 OCT4-ΔPE-GFP+human ESCs derived in 2i/L/DOX. 24h after Dox withdrawal, cells weredissociated in 0.25% trypsin/EDTA solution (Invitrogen) and seeded at adensity of 5000 cells per individual well in 96 well plates on a MEFfeeder layer in 2i/L supplemented with the ROCK-inhibitor Y-27632. Afteran additional 24h, 2i/L medium in each individual 96 well wassupplemented with a kinase inhibitor from the LINCS inhibitor library(Gray Laboratory, Dana Farber Cancer Institute, Boston, Mass.) at afinal concentration of 1 μM. To improve experimental consistency thesmall molecule library was applied using the Caliper RapidPlate 96 wellLiquid Handling System (Zymark, Westborough, Mass.). Briefly, a masterplate containing 10 mM stock solution of the library in DMSO was firstdiluted to 100 μM in HEPES aqueous solution (daughter plate), andsubsequently to 10 μM in N2B27 basal medium supplemented with 2i/L(granddaughter plate). This granddaughter plate was then diluted afurther 10× in 2i/L medium, the final medium was pre-mixed and appliedslowly to the 96 well assay plate. Following two medium changes during aseven day period the proportion of OCT4-ΔPE-GFP+ human ESCs in each wellwas assessed using the High-Throughput System on the LSRFortessa(Beckton-Dickinson, San Jose, Calif.). To screen for small moleculesthat improve the proportion of viable OCT4-ΔPE-GFP+ cells, this assaywas modified by the addition of 1.0 μM SB590885 to each well andinclusion of a 10 minute DAPI (Life Technologies) staining prior tohigh-throughput FACS analysis. Viable cells were gated from theDAPI-negative fraction and subsequently assessed for GFP status.

Immunostaining

Immunostaining was performed according to standard protocols using thefollowing primary antibodies: Oct-3/4 (mouse monoclonal, Santa CruzBiotechnology); hNANOG (Cat. No. AF1997, goat polyclonal, R&D Systems);AFP (Cat. No. A8452, mouse monoclonal, Sigma); HNF4a (goat polyclonal,Santa Cruz); Nestin (Cat. No. AB5922, mouse monoclonal, Milipore); Pax6(Cat. No. PRB-278P, rabbit polyclonal, Covance); appropriate Alexa Fluordye conjugated secondary antibodies (Invitrogen) were used. Nuclei werestained with DAPI (Life Technologies) and analyzed (LSM710, Zeiss;Eclipse Ti—Nikon). Images were taken using LSM710 confocal microscope(Zeiss) or Inverted microscope (Eclipse Ti—Nikon).

RNA FISH and Imaging

RNA FISH was performed as outlined in (Faddah et al., 2013; Raj et al.,2010; Raj et al., 2008). All hybridizations were performed in solutionusing probes coupled to either tetramethylrhodamine (TMR) (Invitrogen),Alexa 594 (Invitrogen) or Cy5 (GE Amersham). TMR was used for the probesagainst human Klf4 mRNA, Alexa 594 for the probes against human Oct4,Rex1, and Nanog mRNA, and Cy5 for the probes against human Oct4 mRNA.Optimal probe concentrations during hybridization were determinedempirically. Imaging involved taking stacks of images spaced 0.4 μmapart using filters appropriate for DAPI, TMR, Alexa 594 and Cy5. Allimages were taken with a Nikon Ti-E inverted fluorescence microscopeequipped with a 100× oil-immersion objective and a Photometrics Pixis1024 CCD camera using MetaMorph software (Molecular Devices, Downington,Pa.). During imaging, photobleaching was minimized through the use of anoxygen-scavenging solution using glucose oxidase. The cells weresegmented manually and counted the number of fluorescent spots, each ofwhich corresponds to an individual mRNA, using a combination of asemi-automated method described in (Itzkovitz et al., 2011; Raj et al.,2008) and custom software written in MATLAB (Mathworks).

RNA Extraction and Synthetic RNA Spike-In

Total RNA and sample preparation was performed as previously described(Loven et al., 2012). Briefly, 1 million naïve or primed human ESCs weretrypsinized and separated from GFP-labeled MEFs using the FACSAria(Beckton-Dickinson) prior to lysis and RNA extraction. Biologicalduplicates were subsequently collected and homogenized in 1 ml of TRIzolReagent (Life Technologies, 15596-026), purified using the mirVANA miRNAisolation kit (Ambion, AM 1560) following the manufacturer'sinstructions and resuspended in 100 ml nuclease-free water (Ambion,AM9938). Total RNA was spiked-in with ERCC RNA Spike-In Mix (Ambion,4456740), treated with DNA-Free™ DNase I (Ambion, AM1906) and analyzedon Agilent 2100 Bioanalyzer for integrity. RNA with the RNA IntegrityNumber (RIN) above 9.8 was hybridized to GeneChip PrimeView Human GeneExpression Arrays (Affymetrix).

Microarray Sample Preparation and Analysis

For microarray analysis, total RNA samples were used for microarrayexpression analysis. 100 ng of total RNA was used to preparebiotinylated cRNA (cRNA) according to the manufacturer's protocol (30IVT Express Kit, Affymetrix 901228). GeneChip arrays (Primeview,Affymetrix 901837) were hybridized and scanned according to standardAffymetrix protocols. The raw data was obtained by using Affymetrix GeneChip Operating Software using default settings. A Primeview CDF providedby Affymetrix was used to generate .CEL files. The CEL files wereprocessed with the expresso command to convert the raw probe intensitiesto probeset expression values with MASS normalization using the standardtools available within the affy package in R. The probesets of the samegene were next collapsed into a single value to represent the gene bytaking the mean value. Differential gene expression was determined usingmoderated t-statistic in the “limma” package(http://bioinf.wehi.edu.au/limma/) from Bioconductor(www.bioconductor.org) (Smyth, 2004). A gene was considereddifferentially expressed if it met the following criteria: 1) absolutelog 2 fold-change ≧1 between the mean expression of the two condition,2) adjusted p-value less than 0.1 by a moderated t-test within the limmapackage with BH multiple hypothesis testing correction.

The previously published expression profiles from the naïve human EScells were also processed (GSM1139484 and GSM1139494) and primed EScells (GSM1139488 and GSM1139495) from Gafni et al., 2013. TheAffymetric CDF file (version V1.r3) was used to generate .CEL files. TheCEL files were processed with RMA normalization using the standard toolsvariable within the affy package in R. The p-values for differentialgene expression analysis were determined using moderated t-statistic inthe “limma” package (http://bioinf.wehi.edu.au/limma/) from Bioconductor(www.bioconductor.org) (Smyth, 2004).

Cross-Species Gene Expression Analysis

Cross-species gene expression analysis was performed as previouslydescribed (Gafni et al., 2013) with some modifications. Previouslypublished mouse ES cell and EpiSC cell gene expression datasets on anAgilent 4×44 k array platform (GSE15603). Probeset mapping between theAgilent 4×44 k array platform and Affymetric Primeview platform ofhuman-mouse homologous genes was downloaded from Enesembl biomart(www.ensembl.org/biomart). The probesets of the same gene in human ormouse were next collapsed into a single value to represent the geneindependently in each species by taking the mean value. The relativeexpression values from mouse and human were next calculatedindependently by divining the expression values of the samples by themean of expression values within the same genes across samples in thesame species. Pair-wise comparisons were performed on the relativeexpression values from the human and mouse expression profiles usingPearson correlation coefficients (PCCs). The average linkagehierarchical clustering of the Pearson correlation was shown in theheatmap.

Chromatin Immunoprecipitation (ChIP)

Cells were crosslinked for 10 minutes at room temperature by theaddition of one-tenth of the volume of 11% formaldehyde solution (11%formaldehyde, 50 mMHEPES pH 7.3, 100 mMNaCl, 1 mMEDTA pH 8.0, 0.5 mMEGTApH 8.0) to the growth media followed by quenching with 100 mM glycine.Cells were washed twice with PBS, then the supernatant was aspirated andthe cell pellet was flash frozen in liquid nitrogen. Frozen crosslinkedcells were stored at −80 C. 20 ul of Dynal magnetic beads (Sigma) wereblocked with 0.5% BSA (w/v) in PBS. Magnetic beads were bound with 2 ugof the indicated antibody. The antibodies used were as follows: H3K4me3(Abcam ab8580) and H3K27me3 (Millipore 07-449). Crosslinked cells werelysed with lysis buffer 1 (50 mMHEPES pH 7.3, 140 mMNaCl, 1 mMEDTA, 10%glycerol, 0.5% NP-40, and 0.25% Triton X-100) and resuspended andsonicated in sonication buffer (50 mM Tris-HCl [pH 7.5], 140 mMNaCl, 1mMEDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS). Cells weresonicated at 4 C with a Bioruptor (Diagenode) at high power for 25cycles for 30 s with 30 s between cycles. Sonicated lysates were clearedand incubated overnight at 4_C with magnetic beads bound with antibodyto enrich for DNA fragments bound by the indicated factor. Beads werewashed two times with sonication buffer, one time with sonication bufferwith 500 mM NaCl, one time with LiCl wash buffer (20 mM Tris pH 8.0, 1mM EDTA, 250 mM LiCl, 0.5% NP-40, 0.5% Na-deoxycholate) and one timewith TE with 50 mM NaCl. DNA was eluted in elution buffer (50 mMTris-HCL pH 8.0, 10 mM EDTA, 1% SDS). Cross-links were reversedovernight. RNA and protein were digested using RNase A and Proteinase K,respectively and DNA was purified with phenol chloroform extraction andethanol precipitation.

Illumina Sequencing and Library Generation

Purified ChIP DNA was used to prepare Illumina multiplexed sequencinglibraries. Libraries for Illumina sequencing were prepared following theIllumina TruSeq DNA Sample Preparation v2 kit protocol with thefollowing exceptions. After end-repair and A-tailing, ImmunoprecipitatedDNA (_10-50 ng) or Whole Cell Extract DNA (50 ng) was ligated to a 1:50dilution of Illumina Adaptor Oligo Mix assigning one of 24 uniqueindexes in the kit to each sample. Following ligation, libraries wereamplified by 18 cycles of PCR using the HiFi NGS Library Amplificationkit from KAPA Biosystems. Amplified libraries were then size-selectedusing a 2% gel cassette in the Pippin Prep system from Sage Science setto capture fragments between 200 and 400 bp. Libraries were quantifiedby qPCR using the KAPA Biosystems Illumina Library Quantification kitaccording to kit protocols. Libraries with distinct TruSeq indexes weremultiplexed by mixing at equimolar ratios and running together in a laneon the Illumina HiSeq 2000 for 40 bases in single read mode.

Gene Sets and Annotations

All analysis was performed using RefSeq (NCBI37/HG19) (Pruitt et al.,2007) human gene annotations.

ChIP-Seq Data Processing

All ChIP-Seq datasets were aligned using Bowtie (version 0.12.9)(Langmead et al., 2009) to build version NCBI37/HG19 of the human genomeusing -n2, -e70, -m2, -k2, -best. The MACS version 1.4.1 (Model basedanalysis of ChlP-Seq) was used (Zhang et al., 2008) peak findingalgorithm to identify regions of ChIP-Seq enrichment over background.

A p-value threshold of enrichment of 1e-9 was used for all datasets.

Heatmap Representation of Read Density Profiles.

A gene was defined as Polycomb-associated if an enriched region forH3K27me3 (representing polycomb complexes) was located within +/−1 kb ofthe TSS. H3K27me3 is a histone modification associated with Polycombcomplexes (Boyer et al., 2006). The annotated TSS of Polycomb-associatedgenes were aligned at the center in the composite view of signal densityprofile. The average ChIP-seq read density (r.p.m./bp) around 5 kbcentered on the centers in 50 bp bin was calculated.

Meta Representations of ChIP-Seq Occupancy

Genome-wide average “meta” representations of ChIP-seq occupancy ofdifferent factors were created by mapping ChIP-seq read density toPolycomb-associated genes. Three sets of regions were created: upstream,gene body and downstream. 80 equally-sized bins divided the −2000 to 0promoter region, 200 equally-sized bins divided the length of the genebody, and 80 equally-sized bins divided the 0 to +2 kb downstreamregion. The average ChIP-Seq factor density in each bin was calculatedto create a meta genome-wide average in units of rpm/bp.

Differentiation Assays Teratoma Formation

Single cell dissociations of naïve human ESCs were resuspended in 250 μlof medium and co-injected subcutaneously with 250 ul of matrigel in theflank of NOD/SCID mice. Tumors generally developed within 8 to 12 weeksand animals were sacrificed before tumor size exceeded 3 cm in diameter.Teratomas were isolated after sacrificing the mice and fixed informalin. After sectioning, teratomas were diagnosed based onhematoxylin and eosin staining.

Directed Differentiation into Hepatocytes

Differentiation of naïve human ESCs into hepatocytes was obtained asdescribed previously in conventional human ESCs (Si-Tayeb et al., 2010).Single cells were cultivated on Matrigel coated plates (2 mg/ml) underlow oxygen conditions. Differentiation was initiated by cultivatingcells for 5 days in Activin A (100 ng/ml) containing RPMI/B27 mediumunder ambient oxygen, followed by 5 days in BMP4 (20 ng/ml)/FGF-2 (10ng/ml) containing RPMI/B27 and 5 days in HGF (20 ng/ml) containingRPMI/B27 under 5% oxygen. Finally cells were cultured for 5 days inHepatocyte Culture Medium supplemented with Oncostatin-M (20 ng/ml)under ambient oxygen conditions. Generated hepatocytes were identifiedby expression of AFP, HNF4a and human Albumin.

Morula and Blastocyst Injection

Six to eight weeks old B6D2F1 females were superovulated with 7.5 I.U.of Pregnant Mare Serum (PMS) each given by intraperitoneal (IP)injections followed by an IP injection of 7.5 I.U. of Human ChorionicGonadotropin (HCG) 46 to 48 hours later. They were then mated withB6D2F1 stud males and checked for copulatory plugs the following day.One-cell fertilized embryos were harvested and incubated at 37° C., 5%O₂ for 2-3 days in KSOM medium (Zenith Biotech). At the 8-cell, morula,and blastocyst stage, embryos were injected with 10-15 naïve human ESCsusing a 16 μm piezo needle (Humagen). During injection, the human cellswere kept continuously in drops of their own culture medium, whereas theembryos were kept in M2+ROCKi drops. After injection, the injectedembryos were cultured in KSOM+ROCKi for 3-4 hours, then washed in 5individual drops of KSOM and cultured overnight to the blastocyst stage.When the injected embryos reached the blastocyst stage, 20 embryos weretransferred into each E2.5 p.c pseudopregnant female by uterinetransfer. Seven to eight days later, the post-op females were sacrificedby CO₂ asphyxiation and embryos were harvested at E9.5-E10.5 p.c foranalysis.

qRT-PCR

Total RNA was isolated using the Rneasy Kit (QIAGEN) and reversedtranscribed using the Superscript III First Strand Synthesis kit(Invitrogen). Quantitative RT-PCR analysis was performed in triplicateusing the ABI 7900 HT system with FAST SYBR Green Master Mix (AppliedBiosystems). Gene expression was normalized to GAPDH. Error barsrepresent the standard deviation (SD) of the mean of triplicatereactions. Primer sequences are included in Table 1.

TABLE 1 Primers used in this study SEQ ID Gene Primer sequence (5′-3′)Application NO: NANOG-F GCAGAAGGCCTCAGCACCTA RT-PCR  1 NANOG-RAGGTTCCCAGTCGGGTTCA  2 OCT4-F GCTCGAGAAGGATGTGGTCC RT-PCR  3 OCT4-RCGTTGTGCATAGTCGCTGCT  4 SOX2-F CACTGCCCCTCTCACACATG RT-PCR  5 SOX2-RTCCCATTTCCCTCGTTTTTCT  6 STELLA-F GTTACTGGGCGGAGTTCGTA RT-PCR  7STELLA-R TGAAGTGGCTTGGTGTCTTG  8 KLF4-F GATGGGGTCTGTGACTGGAT RT-PCR  9KLF4-R CCCCCAACTCACGGATATAA 10 GAPDH-F CGAGATCCCTCCAAAATCAA RT-PCR 11GAPDH-R ATCCACAGTCTTCTGGGTGG 12 REX1-F GGAATGTGGGAAAGCGTTCGT RT-PCR 13REX1-R CCGTGTGGATGCGCACGT 14 PRDM14-F TGAGCCTTCAGGTCACAGAG RT-PCR 15PRDM14-R ATTTCCTATCGCCCTTGTCC 16 EGFP-F AGAACGGCATCAAGGTGAAC RT-PCR 17EGFP-R TGCTCAGGTAGTGGTTGTCG 18 FUW-KLF2-F GATTTTGCTGGGTTGGTTTTT RT-PCR19 FUW-KLF2-R CCACATAGCGTAAAAGGAGCA 20 FUW-NANOG-F GCTGGGGAAGGCCTTAATGTRT-PCR 21 FUW-NANOG-R CCACATAGCGTAAAAGGAGCA 22 PAX6-FCTTTGCTTGGGAAATCCGAG RT-PCR 23 PAX6-R AGCCAGGTTGCGAAGAACTC 24

Results A Reporter System for Naïve Human Pluripotency Based on OCT4Distal Enhancer Activity

An important molecular signature of naïve pluripotency in the mousesystem is the use of the distal enhancer (DE) of OCT4. This elementcontrols Oct4 expression in naïve mouse ESCs, pre-implantation mouseembryos and germ cells (Yeom et al., 1996). In contrast, expression ofOct4 in primed EpiSCs and the mouse post-implantation embryo is undercontrol of the proximal enhancer (PE) element (Tesar et al., 2007). Todetect rare naïve human ESCs in a large population of primed cells, areporter system for OCT4 DE activity was engineered using TALENs. The PEelement was deleted in an OCT4-GFP allele that was previouslyestablished in WIBR3 primed human ESCs (Hockemeyer et al., 2011) (FIG.1A and FIG. 8A). TALENs were designed to cleave in the 5′ end of the PE,together with a donor vector containing LoxP sites bordering aselectable marker and gene sequences homologous to those flanking thePE. After targeting the allele harbors an approximately 1 Kb deletion ofthe PE sequence. Successful integration of this PE targeting vector(FIG. 1B), and subsequent removal of the selection cassette (FIG. 8A)were confirmed. As expected, deletion of the PE resulted in substantialattenuation of the OCT4-GFP signal (FIG. 1C). Single molecule RNA FISHanalysis in individual cells showed that GFP expression was diminished,while OCT4 expression was reduced approximately by 50%, following PEremoval (FIG. 1D). These changes in OCT4 and GFP expression demonstratethat OCT4 expression in primed human ESCs is primarily dependent on thePE, rather than the DE, as observed in mouse EpiSCs.

It was investigated whether overexpression of transcription factorsspecific to naïve pluripotency together with the application ofserum-free 2i/L culture conditions would result in augmentedOCT4-ΔPE-GFP activity. For this reason the transcription factors KLF2and NANOG were overexpressed using doxycycline (DOX) induciblelentiviral expression vectors. Compared to its family members Klf4 andKlf5, Klf2 has enhanced capability to induce naïve pluripotency in mouseEpiSCs (Hall et al., 2009). In addition, the homeodomain transcriptionfactor Nanog is critical for the establishment of naïve pluripotency(Silva et al., 2009) and can revert EpiSCs to the naïve state in theabsence of kinase inhibition (Theunissen et al., 2011). Consistent withthe potent effects of these factors in the mouse system, combinedoverexpression of KLF2 and NANOG in primed human ESCs resulted inincreased OCT4-ΔPE-GFP reporter levels in a fraction of cells (FIG. 1Eand FIG. 8B). The appearance of high OCT4-ΔPE-GFP+ cells was strictlydependent on the expression of both factors, and could only be observedin presence of 2i/L/DOX. GFP+ colonies were clonally expanded on a mouseembryonic fibroblast (MEF) feeder layers in the presence of 2i/L/DOX,while retaining a dome-like colony morphology and pluripotency geneexpression (FIG. 1F and FIG. 1G). Strikingly, withdrawal ofDOX-dependent KLF2 and NANOG expression resulted in the rapid loss ofcolony morphology, appearance of differentiated cells and shutdown ofOCT4-ΔPE-GFP reporter activity within 7 days (FIG. 1F, Right). Thus,2i/L culture conditions are insufficient to maintain the self-renewal ofOCT4-ΔPE-GFP+ cells after withdrawal of exogenous factor expression. Therapid downregulation of naïve reporter activity provides a defined timewindow in which to screen for small molecules that support themaintenance of a putative naïve human pluripotent state.

The inability to maintain OCT4-ΔPE-GFP activity in 2i/L led us tofurther investigate the consequences of these culture conditions whenapplied directly to conventional human ESCs in the absence of transgeneexpression. In the mouse system dual MEK and GSK3 inhibition is thoughtto consolidate the ground state and eliminate EpiSCs and otherdifferentiated phenotypes that cannot survive under these minimalconditions (Silva and Smith, 2008). In contrast, rapid expansion inserum-free 2i/L was observed of initially dome-shaped human ESC coloniesthat assumed a neural morphology upon further passaging (FIG. 1H). Flowcytometric analysis revealed that weak levels of OCT4-ΔPE-GFP detectedin primed human ESCs completely disappeared in 2i/L alone (FIG. 1E).Consistent with the morphological change, the loss of OCT4 and NANOGexpression and upregulation of NESTIN and PAX6 expression in primedhuman ESCs expanded in serum-free 2i/L medium was observed (FIG. 1I andFIG. 8C-D). These observations are in agreement with a report that 2i/Ltreatment induces differentiation of human ESCs into primitive neuralstem cells (Hirano et al., 2012). We conclude that 2i/L conditions donot interfere with expansion of differentiated cell types in human cellsto the same extent as in the mouse system and are consequentlyinsufficient for stabilizing naïve human pluripotency (FIG. 1J).

Identification of Compounds that Maintain Naïve Reporter Activity UponTransgene Withdrawal

To identify compounds that sustain OCT4-ΔPE-GFP activity in the absenceof KLF2 and NANOG expression, a kinase inhibitor library was screened inthe presence of 2i/L for a period of 7 days after DOX withdrawal in 96well format (FIG. 2A). This screen identified 10 different hit compoundsthat partially rescued the proportion of GFP+ cells when assayed byhigh-throughput FACS analysis (FIG. 2B-C and FIG. 9A). Notable amongthese hits were four separate inhibitors of BRAF and four inhibitors ofupstream receptor tyrosine kinases (RTKs), including DDR1, VEGFR1 andFGFR1. A validation experiment was then performed in 6 well format bywithdrawing DOX and culturing the cells in 2i/L supplemented with eachhit compound for up to 10 passages (FIG. 2D). Nearly all hit compoundsmaintained a proportion of GFP+ cells similar to that observed with DOX.However, the BRAF inhibitor SB590885 preserved the best colonymorphology and proliferation. In contrast, treatment with the FGFreceptor inhibitor PD173074 or pan-RTK inhibitor WZ-4-145 resulted inmaintenance of OCT4-ΔPE-GFP activity in colonies with a disorganizedmorphology. Gene expression analysis 5 passages after DOX withdrawalconfirmed the absence of KLF2 and NANOG transgene expression andretention of endogenous OCT4 and GFP transcripts (FIG. 2E and FIG. 9B).To validate these findings in an independent system DOX-inducible KLF2and NANOG transgenes were introduced in a line of wild-type WIBR3 humanESCs. Upon application of DOX and 2i/L, dome-shaped colonies appearedthat could be expanded clonally. After DOX withdrawal these lines weremaintained in 2i/L/SB590885 while retaining both colony morphology andthe expression of endogenous OCT4 (FIG. 2F-H). Thus, high-throughputchemical screening identified a number of kinase inhibitors, mostnotably the BRAF inhibitor SB590885, that synergize with 2i/L tomaintain OCT4-ΔPE-GFP reporter activity and pluripotency gene expressionin human ESCs after removal of exogenous KLF2 and NANOG expression.

Using a FACS-based live/dead discrimination assay it was determined thatcells maintained in 2i/L/SB590885 had reduced viability (FIG. 9C). Thisled us to consider whether other small molecules could cooperate with2i/L/SB590885 to improve the fraction of viable GFP+ cells. Inclusion ofa live/dead assay enhanced the resolution of the 96 well high-throughputFACS analysis, allowing for more sensitive discrimination between theproportion of viable OCT4-ΔPE-GFP+ cells cultured in 2i/L/DOX vs.2i/L/SB590885 (FIG. 3A). This viability assay was included in a modifiedscreen in which GFP+ cells were cultured for two passages after DOXwithdrawal in 2i/L/SB590885 supplemented with the kinase inhibitorlibrary (FIG. 3B-C and FIG. 10A). This screen identified several hitcompounds that improved the fraction of viable GFP+ cells, including theLCK/SRC inhibitor WH-4-023 (FIG. 10B). A titration of the concentrationsof PD0325901, CHIR99021 and SB590885 was also performed during twopassages after DOX withdrawal followed by high-throughput FACS analysis(FIG. 3D). A significant improvement was observed in the proportion ofviable GFP+ cells using a concentration of 1 M PD0325901, 0.3 μMCHIR99021 and 0.5 μM SB590885. Notably, lower concentrations ofCHIR99021 improved the proportion of viable GFP+ cells, whereas higherconcentrations of CHIR99021 had an opposite effect. This is reminiscentof recent evidence that lowered GSK3 inhibition reduces differentiationand enhances the self-renewal of rat ESCs (Meek et al., 2013).

The proportion of viable OCT4-ΔPE-GFP+ cells after DOX withdrawal wasfurther improved by combining optimized concentrations of PD0325901,CHIR99021 and SB590885 with 1 μM WH-4-023 (FIG. 3F). The ROCK inhibitorY-27632, which improved the proportion of viable GFP+ cells, was alsoincluded (FIG. 10C). Finally, improved morphology upon long-term culturewas observed without DOX by replacing CHIR99021 with an alternative GSK3inhibitor, IM12 (FIG. 3G and FIG. 10D). Pluripotency of putative naïvehuman ESCs maintained under these conditions was confirmed by generationof high-grade teratomas including tissues representing all three germlayers after subcutaneous injection in NOD/SCID mice (FIG. 3H). Insummary, a combination of five compounds have been provided, includinginhibitors of MEK, GSK3, BRAF, ROCK and SRC, that supports the expansionof viable OCT4-ΔPE-GFP+ human pluripotent cells after removal ofexogenous transcription factor expression.

Derivation of Genetically Unmodified Naïve Human ESCs in 5i/L

The consequences of applying the optimized 5i/L medium to conventionalhuman ESCs was investigated in the absence of ectopic factor expression.Not surprisingly, this highly selective inhibitor cocktail generatedwidespread cell death within 2 days of treatment. However, the emergenceof a small number of dome-shaped colonies was observed within 10 daysthat were positive for the OCT4-ΔPE-GFP reporter allele (FIG. 4B). Inaddition, these colonies were efficiently isolated and clonally expandedafter dissociation in trypsin (FIG. 4B). The appearance of theseOCT4-ΔPE-GFP+ cells using the optimized chemical conditions suggestedthat genetic manipulation, such as overexpression of KLF2 and NANOG, maybe dispensable for driving primed human ESCs to the naïve state.However, the slow and inefficient kinetics of this chemical conversionevent led us to consider whether providing additional growth factorsupport might boost the efficiency of naïve cell induction. Provision of5i/L supplemented with FGF and Activin A (5i/L/FA) enhanced the kineticsof OCT4-ΔPE-GFP induction (FIG. 4C), and enabled conversion of wild-typeWIBR2 primed human ESCs into a cell state with identical morphology andnormal karyotype (FIG. 4D and FIG. 11A). Given the selective nature ofthe 5i culture regimen, it is speculated that additional growth factorsupport prolongs the time window during which primed human ESCs areamenable to convert to the naïve state. Consideration was taken towardculture conditions and if they would support the direct derivation ofnovel ESC lines from human blastocysts. Application of 5i/L/FA to humanblastocyst outgrowths resulted in the establishment of a human ESC linewith a similar dome-shaped colony morphology and normal karyotype (FIG.4E and FIG. 11B). Hence, 5i/L/FA promotes induction of OCT4-ΔPE-GFPactivity in the absence of reprogramming transgenes, and the derivationof putative naïve human ESCs directly from blastocysts.

To investigate the relative contribution of individual components in5i/L/FA medium, single kinase inhibitors or growth factors were removedfrom a clonal line of OCT4-ΔPE-GFP+ cells derived in the absence oftransgenes. Withdrawal of the MEK inhibitor PD0325901 or BRAF inhibitorSB590885 resulted in the rapid and widespread loss of colony morphology(FIG. 11C), OCT4-ΔPE-GFP reporter activity (FIG. 4F) and pluripotencygene expression (FIG. 4G). Withdrawal of the SRC inhibitor WH-4-023caused a change in morphology (FIG. 11C) and slight reduction inOCT4-ΔPE-GFP reporter activity (FIG. 4F). In addition, withdrawal of theROCK inhibitor Y-27632 caused a significant reduction in proliferation(FIG. 11C). On the other hand, withdrawal of FGF had no apparent effectby any of the parameters examined, which is consistent with this growthfactor being involved in the maintenance of primed pluripotency.Similarly, withdrawal of Activin A did not cause a reduction inOCT4-ΔPE-GFP activity (FIG. 4F). However, more differentiation andreduced expression was observed of pluripotency genes when FGF andActivin A were removed together (FIG. 4G and FIG. 11C). Surprisingly,reporter activity and pluripotency gene expression were unaffected bythe removal of either GSK3 inhibition or hLIF (FIG. 4F-G). Therefore,the maintenance of OCT4-ΔPE-GFP reporter activity is dependent primarilyon MEK inhibition and BRAF inhibition, while robust proliferation ofGFP+ cells requires ROCK inhibition. Recombinant FGF enhances theinduction of naïve reporter activity, but can be omitted in establishedGFP+ cells.

Evaluation of Alternative Culture Systems for Naïve Human Pluripotency

Recently, several groups reported alternative conditions for inducing anaïve pluripotent state in conventional human ESCs (Chan et al., 2013;Gafni et al., 2013; Valamehr et al., 2014; Ware et al., 2014).Comparison of the culture components in these studies with the mediashows both commonalities and differences (FIG. 5A). All previouslypublished protocols for naïve human pluripotency include 2i. Anotherubiquitous component is FGF, which is added either as a recombinantprotein (Gafni et al., 2013; Valamehr et al., 2014; Ware et al., 2014)or present at high levels in mTesr basal medium (Chan et al., 2013).However, the use of additional FGF-Raf-Erk inhibitors, the primary hitsfrom the kinase inhibitor screen, was not previously reported.Furthermore, while these studies considered various criteria fordefining naïve human pluripotency, endogenous OCT4 distal enhanceractivity was not assessed. Therefore, it was examined whether previouslypublished culture conditions for naïve human pluripotency could activatethe reporter system.

Remarkably, increased levels of OCT4-ΔPE-GFP activity were exclusivelyobserved upon application of 5i/L/A (FIG. 5B). Whereas the naïveconditions described in Gafni et al. (2013) were capable of maintainingregular OCT4-GFP reporter activity after removal of KLF2 and NANOGexpression, these conditions did not maintain OCT4-ΔPE-GFP activity(FIG. 5C). This result is consistent with the observation that none ofsix JNK inhibitors and seven p38 MAP kinase inhibitors present in thekinase inhibitor library showed ability to maintain OCT4-ΔPE-GFPactivity after withdrawal of KLF2 and NANOG expression (FIG. 2B and FIG.9A). These findings suggest that the combination of kinase inhibitorsinduces a novel and distinct state of human pluripotency.

The kinase inhibitors reported by Hanna and colleagues may have anadditive effect in combination with the 5i/L/A medium. Inclusion of theJNK inhibitor SP600125 and/or p38 MAP kinase inhibitor BIR^(B796) didnot affect the proportion of OCT4-ΔPE-GFP+ cells (FIG. 12A). However, anincrease in expression of KLF4 and KLF2 was observed upon addition ofSP600125 (FIG. 12B). A beneficial effect was not observed when addinginsulin (FIG. 12A and FIG. 5C). Another difference between the naïvehuman studies reported to date is the use of 20% knock-out serumreplacement (KSR) or Albumax-containing medium (Gafni et al., 2013;Valamehr et al., 2014; Ware et al., 2014) vs. serum-free N2B27 medium(this study). The consequences of applying the inhibitor cocktail in thepresence of 20% KSR was therefore investigated. Remarkably, this switchin basal medium resulted in the rapid attenuation of OCT4-ΔPE-GFP signalconcomitant with morphological changes (FIG. 5D and FIG. 12C). Hence, ahigh concentration of KSR appears to be detrimental to naïve reporteractivity, independent of additional kinase inhibition. In fact, areduction in pluripotency gene expression was observed upon provision ofKSR or FBS at concentrations >5% (FIG. 5E). However, including 0.5-1%KSR in combination with 5i/L, JNKi and Activin A (6i/L/A) enhanced theefficiency of OCT4-ΔPE-GFP induction from the primed state (FIG. 5F). Itis concluded that the induction and maintenance of OCT4-ΔPE-GFP+ humanESCs is highly sensitive to the choice of basal medium.

Molecular Characterization of Naïve Human Pluripotency

To characterize the gene expression profile of naïve human ESCs derivedunder the conditions, RNA was collected from WIBR2 and WIBR3 human ESCscultured in primed medium or 6i/L/A, and embryo-derived naïve WIN1 cellscultured in 5i/L/A or 6i/L/A. Expression analysis on Affymetrix arrayswere then performed using RNA spike-in normalization (Loven et al.,2012). Cross-species gene expression comparison demonstrated that primedWIBR2 and WIBR3 human ESCs clustered with primed mouse EpiSCs, whilenaïve human ESCs cultured in 5i/L/A or 6i/L/A clustered with naïve mouseESCs (FIG. 6A). The naïve human ESC samples clustered together closely,and addition of the JNK inhibitor had little impact on overall geneexpression in the naïve state (FIG. 6A). The most upregulated geneontology (GO) categories in the naïve state were associated withtranscriptional control, while the most down-regulated categories wereimplicated in neural differentiation and cell adhesion (FIG. 6B).Comparison with the gene expression data published by Gafni et al.(2013) indicated that the conditions for naïve human pluripotency inducegreater transcriptional differences with the primed state (FIG. 6C). Inparticular, 3238 differentially expressed genes (DEGs) were observedcompared to primed human ESCs, whereas Gafni et al. reported 831 DEGs(where log₂ fold change >1 and <−1 & p<0.05).

Intriguingly, the naïve human ESCs exhibited marked downregulation ofthe transcription factors OTX2 and ZIC2/3, which were recently shown todirect Oct4 to primed state-specific enhancer sites in mouse EpiSCs(Buecker et al., 2014). In contrast, these transcription factors wereslightly upregulated in the naïve human ESCs reported by Gafni et al.(2013). The list of differentially expressed genes was examined forpresence of bona fide markers of naïve pluripotency. A number oftranscription factors typically associated with the self-renewal andpluripotency of mouse ESCs ranked among the most highly upregulatedgenes in 5i/L/A or 6i/L/A, including DPPA5, DPPA3 (also known asSTELLA), DPPA2, REX1, KLF4, KLF5, TFCP2L1, and NANOG (FIG. 6D-E).Expression of these factors was largely unaffected in the conditions fornaïve human pluripotency described by Gafni et al. (2013) (FIG. 6D).

Upregulation of transcripts specific to naïve pluripotency under theconditions was confirmed by qRT-PCR (FIG. 6F and FIG. 13A). A recentsingle cell RNA-Seq analysis revealed that these markers of naïvepluripotency were highly upregulated at the morula and epiblast stagesof human pre-implantation development compared to conventional (primed)human ESCs (Yan et al., 2013) (FIG. 13B). This suggests that 5i/L/Aculture re-establishes an early pre-implantation epiblast-specific geneexpression signature that is lost during derivation of human ESCs underconventional conditions. Hence, the conditions induce a uniquetranscriptional profile in human ESCs, characterized by upregulation ofnaïve-specific transcription factors and suppression of neuraldifferentiation genes.

A defining feature of ground state pluripotency in the mouse system isthat transcriptional regulators such as Nanog are expressedhomogeneously (Wray et al., 2010). In contrast, human ESCs underconventional conditions are known to exhibit significant variability ingene expression at the single cell level (Hough et al., 2009). It wastherefore investigated whether human ESCs cultured under the conditionsare more homogeneous with respect to expression of NANOG by singlemolecule (sm) RNA FISH analysis. The mean number of OCT4 mRNAs per cellwas approximately similar between WIBR2 human ESCs in primed medium, thenaïve medium of Gafni et al. (2013), and 5i/L/A (FIG. 6G). As expectedfrom array and qRT-PCR analyses (FIG. 6D and FIG. 6F), the mean numberof NANOG mRNAs per cell was significantly higher in 5i/L/A (FIG. 6G-H).Intriguingly, 5i/L/A culture also resulted in reduced cell-to-cellvariability in NANOG expression (FIG. 13C). Thus, the increasedexpression level of NANOG in 5i/L/A does not arise from a subset ofcells, but is uniform across the population. It was also confirmed byRNA FISH that single cells cultured in 5i/L/A express significantlyhigher numbers of KLF4 and REX1 mRNAs (FIG. 6H).

Chromatin immunoprecipitation was performed followed by DNA sequencing(ChIP-Seq) analysis to determine the genome-wide distribution of theactivation-associated marker, trimethylation of histone 3 lysine 4(H3K4me3) and the transcriptional-silencing-associated marker,trimethylation of histone 3 lysine 27 (H3K27me3). Developmental geneswith bivalent domains marked by the presence of both H3K4me3 andH3K27me3 in the primed state, such as HOXA9, FOXA2, GATA6 and NKX2.5,exhibited a reduced H3K27me3 signal in the naïve state (FIG. 7A). Thehistone methylation profile at loci encoding naïve-specific pluripotencyregulators was then examined. KLF2, KLF4 and KLF5 were bivalent in theprimed state, but almost entirely lost the H3K27me3 mark in the naïvestate (FIG. 7B). Other markers of ground state pluripotency that arehighly upregulated in the system, such as DPPA5, DPPA3 and REX1,acquired H3K4me3 signal during conversion from primed to naïvepluripotency (FIG. 7C). The core pluripotency determinants OCT4, SOX2and NANOG were marked exclusively by H3K4me3 in naïve and primed humanESCs (FIG. 7D). However, a slightly higher H3K4me3 signal was observedat the NANOG promoter in the naïve state, consistent with increasedtranscription (FIG. 6A). Finally, while H3K27me3 was generally depletedfrom promoter regions in the naïve state, some genes that were stronglydown-regulated acquired H3K27me3 signal (FIG. 7E). Consistent withobservations in naïve pluripotent mouse ESCs (Marks et al., 2012),H3K27me3 was strongly reduced at the transcriptional start site (TSS) ofPolycomb group target genes in human ESCs cultured in 6i/L/A (FIG.7F-G). It is concluded that the conversion from primed to naïve humanpluripotency is accompanied by the dynamic rearrangement of activatingand repressive histone modifications. In particular, transcriptionalupregulation of specific regulators of ground state pluripotency isassociated with the gain of H3K4me3 or loss of H3K27me3 in alocus-specific manner.

Developmental Potential of Human ESCs in 5i/L/a

The classical assay for pluripotency of human ESCs and iPSCs is toexamine teratomas formed after subcutaneous injection in NOD/SCID mice.Naïve human ESCs converted under the optimized conditions contributed tohigh-grade teratomas containing tissues representing all three germlayers, regardless of the presence of FGF and Activin in the medium(FIG. 14A). As the practical utility of human pluripotent stem cellsresides mainly in the capacity to differentiate into defined lineages invitro, the suitability of naïve human ESCs in directed differentiationwas also investigated. For this purpose human ESCs derived from WIBR2and WIBR3 primed cells in 5i/L/A were subjected to a five-step protocolfor hepatic differentiation (Si-Tayeb et al., 2010). Hepatocytespositive for the mature markers AFP and HNF4a were generated from bothlines of naïve ESCs (FIG. 14B).

In the mouse system, naïve ESCs are functionally distinguished by thecapability to colonize the embryo and contribute to chimeric animals. Incontrast, primed EpiSCs contribute only very inefficiently to chimeras(Brons et al., 2007; Tesar et al., 2007).

Hanna and colleagues reported that human ESCs cultured in the presenceof 2i, hLIF, JNK inhibition, p38 inhibition, FGF and TGFβ contributedrobustly to interspecies chimeric embryos after injection into mousemorulae (Gafni et al., 2013). This assay would provide a definingfunctional read-out for naïve human pluripotency. Therefore, areproduction this experiment was attempted using human ESCs containingconstitutive GFP and tdTomato transgenes in the AAVS1 locus. In total,860 embryos (8-cell, morula and blastocyst stages) were injected withGFP and tdTomato-labeled human ESCs cultured in 5i/L/A. 436 embryos werealso injected with C1-AAVS1-GFP cells provided by the Hanna laboratory,which were cultured using the conditions reported by Gafni et al.(2013). Only a fraction of embryos (43-45%) were recovered from bothexperimental groups at E10.5, suggesting that the majority of embryoshad been reabsorbed. No fluorescent signal was detected by microscopy inany embryos recovered at E10.5 (FIG. 14C). Thus, contribution of humanESCs in 5i/L/A or the medium of Gafni et al. (2013) to interspecieschimeras is currently too inefficient for detection by standard visualinspection. The development of novel optical clearing methods mayfacilitate the investigation of low-contribution interspecies chimerismin the future.

The proliferation of naïve human embryonic stem cells was found to beenhanced by the reduction or removal of GSK3 inhibition. Atransgene-dependent naïve human embryonic stem (ES) cell line, asubclone of WIBR3, was used to titrate different concentrations ofinhibitors in 5i/L/A (see Theunissen et al., Cell Stem Cell, 2014). Thecell line used is dependent on Doxycycline (DOX) to maintain theexpression of two lentiviral transgenes, KLF2 and NANOG. Flow cytometryrevealed that proportion of cells positive for the OCT4-ΔPE-GFP reporterin 2i/L/DOX was approximately 60% (FIG. 15A). The cell line waskaryotyped, and showed a normal (46, XX) karyotype. A titration assaywas performed to determine the optimal concentrations of small moleculeinhibitors to maintain naïve human DES cells. The cells were subject tosequential passaging by single cell dissociation at low density (1:10)under four conditions: 2i/L, 5i/L/A, t5i/L/A (0.2 μM GSK3 inhibitorIM12) and 4i/L/A (removal of GSK3 inhibitor IM12). Flow cytometrydemonstrated that the two latter conditions showed higher proportions ofcells positive for the OCT4-ΔPE-GFP reporter (FIG. 15B). QuantitativeRT-PCR was performed, and it confirmed the downregulation of exogenousKLF2 and NANOG transgenes and the primed marker VIMENTIN, as well as theupregulation of naïve markers, including KLF4, REX1 and STELLA int5i/L/A and 4i/L/A (FIG. 15C).

The proliferation of naïve human ES cells in the presence of differentWnt signal modulators was quantified after the withdrawal of DOX fromthe transgene-dependent cell line described above. In a six-well plate,1×10⁵ cells were seeded per well and cells were dissociated andre-seeded at a 1:5 density for three successive passages. Cell numbersat successive passages were recorded under six conditions (n=2 pergroup): 2i/L/DOX (control), 2i/L, 5i/L/A, 4i/L/A (removal of GSK3inhibitor IM12), 4i/L/A+CHIR99021 (1 μM), and 4i/L/A+IWR1 (2.5 μM). Theproliferation was significantly elevated in 4i/L/A compared to 5i/L/A atpassages 2 and 3 (FIG. 16). Replacing IM12 with an alternative GSK3inhibitor, CHIR99021, or addition of the Wnt inhibitor, IWR1, did notfurther stimulate the proliferation of naïve human cells, demonstratingthat the reduction or removal of GSK3 inhibitors further enhances humanES cell proliferation.

The induction of naïve human pluripotency in 5i/L/A or 4i/L/A isassociated with X chromosome reactivation. Using TALEN-mediatedtargeting of both alleles of the X-linked MECP2 gene, a fluorescentreporter system for the X chromosome status of human ES cells wasestablished (FIG. 17A). Conversion of the naïve state in 5i/L/A or4i/L/A (-IM12) results in activation of the tdTomato-labeled allele andGFP activity is maintained, regardless of whether the starting color istdTomato-positive (FIG. 17C) or GFP-positive (FIG. 17B). Therefore, theinduction of naïve human pluripotency results from a switch towardbiallelic expression of X-linked genes.

Discussion

The morphological, molecular and functional similarity between humanESCs and mouse post-implantation epiblast-derived EpiSCs has promptedwidespread interest in capturing the equivalent of a naïve pluripotentstem cell in humans. At the outset of this study, it was considered thatthe identification of putative naïve human ESCs would be greatlyfacilitated by the availability of a selective reporter system. In themouse, enhancer-specific regulation of Oct4 expression is a definingmolecular distinction between ESCs and EpiSCs in vitro (Brons et al.,2007; Tesar et al., 2007), and between the ICM and post-implantationepiblast in vivo (Yeom et al., 1996). The primed-specific PE from anOCT4-GFP allele were deleted using TALEN-mediated genome editing inhuman ESCs (Hockemeyer et al., 2011). The observation that OCT4-GFPexpression was down-regulated following PE removal demonstrated thatthis reporter behaves as expected in the primed state. Conversely,OCT4-ΔPE-GFP activity was strongly induced by combined overexpression ofKLF2 and NANOG. A systematic approach was then taken to screen a diversecollection of 230 kinase inhibitors for the capacity to maintainOCT4-ΔPE-GFP activity after removal of ectopic KLF2 and NANOGexpression. Through iterative screening a combination of five kinaseinhibitors were identified that maintained viable GFP-positive cellsupon transgene withdrawal. Moreover, this kinase inhibitor cocktail wascapable of inducing OCT4-ΔPE-GFP activity when applied directly toconventional (primed) human ESCs in the complete absence ofreprogramming factors.

Previous studies describing the isolation of naïve human ESCs alsoreported transgene-free interconversion from primed to naïvepluripotency (Chan et al., 2013; Gafni et al., 2013; Valamehr et al.,2014; Ware et al., 2014). However, these published protocols did notinduce OCT4-ΔPE-GFP activity. This finding was surprising given thatGafni et al. (2013) reported activation of luciferase and BAC reporterconstructs under the control of the DE of human OCT4, while Ware et al.(2014) observed increased DNasel hypersensitivity at the DE in humanESCs derived in 2i and FGF. Thus, the activation of the DE of endogenousOCT4 may represent a more stringent criterion for the naïve pluripotentstate. A difference in the kinetics of conversion was noticed usingpreviously reported protocols: whereas the emergence of naïve coloniesunder the conditions is a relatively protracted process (ca. 10 days)that occurs after widespread cell death, the application of previouslydescribed naïve conditions was accompanied with little cell death andrapid expansion in primed human ESCs. The slow kinetics of conversionobserved in 5i/L/A are consistent with a reprogramming event towards anovel cellular state, rather than adaptation to an identity closer toconventional (primed) human ESCs. Thus, the observations suggest thatthe chemical screens used in this study have identified a distinct andnovel state of human pluripotency.

There are several important differences in the selection of inhibitors,growth factors and basal medium between this protocol and previouslyreported protocols for capturing naïve human pluripotency. First, theprimary screen for maintenance of OCT4-ΔPE-GFP activity identified anumber of inhibitors of BRAF and upstream RTKs, including VEGFR1/2,FGFR1 and EGFR. These targets were not previously implicated in theestablishment of naïve human pluripotency. The requirement for furtherinhibition of FGF-Ras-MEK signaling contrasts with the dependence on FGFsignaling reported in the published protocols. In the mouse system,independence of FGF signaling is considered to be a hallmark of groundstate pluripotent cells, which are primed for commitment byauto-inductive Fgf4/Erk signaling (Silva and Smith, 2008). Whilerecombinant FGF facilitated the conversion of primed human ESCs intoOCT4-ΔPE-GFP-positive cells, the removal of FGF did not affect reporteractivity or pluripotency gene expression in establishedOCT4-ΔPE-GFP-positive cells.

Unlike previous approaches for capturing transgene-independent naïvehuman ESCs, the protocol uses serum-free N2B27 basal medium. This basalmedium was originally used in neural differentiation, but wassubsequently adapted by the Smith laboratory to develop fully definedconditions for self-renewal of mouse ESCs in combination with LIF andBMP4 (Ying et al., 2003) or 2i (Ying et al., 2008). Maintenance ofOCT4-ΔPE-GFP activity is highly sensitive to the choice of basal medium:addition of as little as 5% FBS or KSR resulted in attenuation of theGFP signal concomitant with a reduction in pluripotency gene expression.However, a low percentage (<1%) of KSR improved the kinetics of inducingGFP-positive cells from the primed state. Thus, previously proposedbasal media for naïve human ESCs do not appear to support themaintenance of OCT4-ΔPE-GFP reporter activity.

Transcriptional profiling of human ESCs cultured in 5i/L/A and primedhuman ESCs indicated that the conditions induce a dramatic upregulationin the expression of transcription factors typically associated withnaïve pluripotency, suggesting that induction of endogenous OCT4-ΔPE-GFPactivity correlates with a broader expression signature of the naïvestate. Intriguingly, many of the top-ranked differentially expressedgenes were also enriched during human pre-implantation development,specifically at the morula and epiblast stages, in comparison toconventional human ESCs (Yan et al., 2013). These transcripts includeseveral factors that possess the capability to instate the naïvepluripotent program in mouse EpiSCs, such as KLF4 (Guo et al., 2009),NANOG (Silva et al., 2009), and TFCP2L1 (Martello et al., 2013; Ye etal., 2013). The latter gene is of particular interest as it wasidentified as the main effector downstream of LIF-STAT3 signaling innaïve mouse ESCs. Induction of TFCP2L1 provides a rationale for thesurprising observation that withdrawal of hLIF does not affectOCT4-ΔPE-GFP activity or pluripotency gene expression in the naïve humanESCs. Strong upregulation of REX1 and STELLA were also observed, both ofwhich are used as reporters of ground state pluripotency in the mousesystem (Hayashi et al., 2008; Marks et al., 2012). Of further interestis that the conditions for naïve human pluripotency induced significantdownregulation of OTX2 and ZIC2/3, which drive primed-specific enhancerbinding of Oct4 in mouse EpiSCs (Buecker et al., 2013). In contrast,these transcription factors were upregulated in the naïve human ESCsreported by Gafni et al. (2013), while typical markers of naïvepluripotency such as NANOG, KLF2, KLF4, STELLA, REX1 and TFCP2L1 werenot induced.

Human ESCs can be expanded in the presence of 2i and various additionalinhibitors and growth factors while maintaining a dome-shaped morphologyand expression of some pluripotency genes (Gafni et al., 2013; Valamehret al., 2014; Ware et al., 2014). These protocols offer practicaladvantages, including enhanced proliferation and single cell cloning,which benefit applications such as gene targeting. As such, thesestudies facilitate the application of human iPSCs in disease modelingand regenerative medicine. The state of pluripotency defined here isbased on activation of the endogenous OCT4-ΔPE-GFP allele along with atranscriptional and epigenomic profile that closely resembles naïvemouse pluripotency, and is distinct from previously reported naïve humanESCs.

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EQUIVALENTS AND SCOPE

In the claims articles such as “a,” “an,” and “the” may mean one or morethan one unless indicated to the contrary or otherwise evident from thecontext. Claims or descriptions that include “or” between one or moremembers of a group are considered satisfied if one, more than one, orall of the group members are present in, employed in, or otherwiserelevant to a given product or process unless indicated to the contraryor otherwise evident from the context. The invention includesembodiments in which exactly one member of the group is present in,employed in, or otherwise relevant to a given product or process. Theinvention includes embodiments in which more than one, or all of thegroup members are present in, employed in, or otherwise relevant to agiven product or process.

Furthermore, the invention encompasses all variations, combinations, andpermutations in which one or more limitations, elements, clauses, anddescriptive terms from one or more of the listed claims is introducedinto another claim. For example, any claim that is dependent on anotherclaim can be modified to include one or more limitations found in anyother claim that is dependent on the same base claim. Where elements arepresented as lists, e.g., in Markush group format, each subgroup of theelements is also disclosed, and any element(s) can be removed from thegroup. It should it be understood that, in general, where the invention,or aspects of the invention, is/are referred to as comprising particularelements and/or features, certain embodiments of the invention oraspects of the invention consist, or consist essentially of, suchelements and/or features. For purposes of simplicity, those embodimentshave not been specifically set forth in haec verba herein. It is alsonoted that the terms “comprising” and “containing” are intended to beopen and permits the inclusion of additional elements or steps. Whereranges are given, endpoints are included. Furthermore, unless otherwiseindicated or otherwise evident from the context and understanding of oneof ordinary skill in the art, values that are expressed as ranges canassume any specific value or sub-range within the stated ranges indifferent embodiments of the invention, to the tenth of the unit of thelower limit of the range, unless the context clearly dictates otherwise.

This application refers to various issued patents, published patentapplications, journal articles, and other publications, all of which areincorporated herein by reference. If there is a conflict between any ofthe incorporated references and the instant specification, thespecification shall control. In addition, any particular embodiment ofthe present invention that falls within the prior art may be explicitlyexcluded from any one or more of the claims. Because such embodimentsare deemed to be known to one of ordinary skill in the art, they may beexcluded even if the exclusion is not set forth explicitly herein. Anyparticular embodiment of the invention can be excluded from any claim,for any reason, whether or not related to the existence of prior art.

Those skilled in the art will recognize or be able to ascertain using nomore than routine experimentation many equivalents to the specificembodiments described herein.

The scope of the present embodiments described herein is not intended tobe limited to the above Description, but rather is as set forth in theappended claims. Those of ordinary skill in the art will appreciate thatvarious changes and modifications to this description may be madewithout departing from the spirit or scope of the present invention, asdefined in the following claims.

What is claimed is:
 1. A method for changing the pluripotency state of avertebrate cell to a more naïve state, the method comprising: culturinga pluripotent vertebrate cell in the presence of aserine/threonine-protein kinase B-Raf (BRAF) inhibitor, an epidermalgrowth factor receptor (EGFR) inhibitor, a vascular endothelial growthfactor 1 (VEGFR1) inhibitor, or a fibroblast growth factor receptor 1(FGFR1) inhibitor; and maintaining the cell in culture under conditionssuitable and a time sufficient to convert the pluripotency state of thevertebrate cell to a more naïve state than the pluripotency state of thevertebrate cell of culturing step.
 2. The method of claim 1, wherein thecell is cultured and maintained in the presence of a mitogen-activatedprotein kinase kinase (MEK) inhibitor.
 3. The method of claims 1 or 2,wherein the cell is cultured and maintained in the presence of aglycogen synthase kinase 3 (GSK3) inhibitor, a rho-associated proteinkinase (ROCK) inhibitor, and/or a proto-oncogene tyrosine-protein kinase(Src) inhibitor.
 4. The method of any one of claims 1-3, wherein thecell is cultured and maintained in the presence of a BRAF inhibitor, aMEK inhibitor, and a GSK3 inhibitor.
 5. The method of any one of claims1-3, wherein the cell is cultured and maintained in the presence of aBRAF inhibitor, a MEK inhibitor, a GSK3 inhibitor, and a ROCK inhibitor.6. The method of any one of claims 1-3, wherein the cell is cultured andmaintained in the presence of a BRAF inhibitor, a MEK inhibitor, a GSK3inhibitor, a ROCK inhibitor, and an Src inhibitor.
 7. The method any oneof claims 1-3, wherein the cell is cultured and maintained in thepresence of a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, and anSrc inhibitor.
 8. A method for changing the pluripotency state of avertebrate cell to a more naïve state, the method comprising: culturinga pluripotent vertebrate cell in the presence of a compound of Formula(A):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of culturing step;wherein: L^(A1) is a substituted or unsubstituted, saturated orunsaturated, C₁₋₄ aliphatic chain, optionally wherein one, two, or threechain atoms of the aliphatic chain are independently replaced with —O—,—S—, —NR^(A5)—, —N═, or ═N—, wherein each instance of R^(A5) isindependently hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; L^(A2) is a substituted or unsubstituted,saturated or unsaturated, C₁₋₆ aliphatic chain, optionally wherein one,two, or three chain atoms of the aliphatic chain are independentlyreplaced with —O—, —S—, —NR^(A6)—, —N═, or ═N—, wherein each instance ofR^(A6) is independently hydrogen, substituted or unsubstituted C₁₋₆alkyl, or a nitrogen protecting group; R^(A1) is hydrogen, substitutedor unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group; eachinstance of R^(A2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; each instance of R^(a)is independently hydrogen, substituted or unsubstituted acyl,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; n2 is 0, 1, 2, 3, 4, 5,6, or 7; each instance of R^(A3) is independently halogen, substitutedor unsubstituted alkyl, substituted or unsubstituted alkenyl,substituted or unsubstituted alkynyl, substituted or unsubstitutedcarbocyclyl, substituted or unsubstituted heterocyclyl, substituted orunsubstituted aryl, substituted or unsubstituted heteroaryl, —OR^(a),—N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; n3 is 0, 1, 2, or 3;each instance of R^(A4) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and n4 is 0, 1, 2, 3,or
 4. 9. The method of claim 8, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 10. The method of claim8, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 11. The method of claim8, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 12. The method of any oneof any one of claims 8-11, wherein R^(A1) is hydrogen.
 13. The method ofany one of claims 8-12, wherein at least one instance of R^(A2) issubstituted or unsubstituted alkyl.
 14. The method of any one of claims8-13, wherein n3 is
 0. 15. The method of any one of claims 8-14, whereinn4 is
 0. 16. The method of any one of claims 8-15, wherein at least oneinstance of R^(A5) is hydrogen.
 17. The method of any one of claims8-16, wherein at least one instance of R^(A6) is hydrogen.
 18. Themethod of claim 8, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 19. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (B):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: L^(B1) is —N(R^(B4))C(═O)—, —C(═O)N(R^(B4))—, or—N(R^(B4))C(═O)N(R^(B4))—, wherein each instance of R^(B4) isindependently hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; when L^(B1) is —N(R^(B4))C(═O)— or—C(═O)N(R^(B4))—, L^(B1) is directly attached to the carbon atom labeledwith 3 or 5; when L^(B1) is —N(R^(B4))C(═O)N(R^(B4))—, L^(B1) isdirectly attached to the carbon atom labeled with 4; L^(B2) is —O—, —S—,—NR^(B5)—, or —C(R^(B6))₂—, wherein R^(B5) is hydrogen, substituted orunsubstituted C₁₋₆ alkyl, or a nitrogen protecting group; and eachinstance of R^(B6) is independently hydrogen, halogen, or substituted orunsubstituted alkyl; each of X^(B1), X^(B2), and X^(B3) is independentlyN or CR^(B7), wherein each instance of R^(B7) is independently hydrogen,halogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two R^(a)groups are joined to form a substituted or unsubstituted heterocyclic orsubstituted or unsubstituted heteroaryl ring; each instance of R^(B1) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a),—C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; o1 is 0, 1, 2, 3, 4, or 5; each instance of R^(B2)is independently halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; o2 is 0, 1, 2, 3, or 4; each instance of R^(B3) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂,or an instance of R^(B3) and an instance of R^(B7) are joined to form asubstituted or unsubstituted heterocyclic ring; and o3 is 0, 1, or 2.20. The method of claim 19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 21. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 22. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 23. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein each of R^(B9)and R^(B10) is independently substituted or unsubstituted, monocyclic,5- to 6-membered heteroaryl, wherein one, two, three, or four atoms inthe heteroaryl ring system are independently nitrogen, oxygen, orsulfur.
 24. The method of claim 19, wherein the compound is of theformula:

or a pharmaceutically acceptable salt thereof.
 25. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein Ring B is asubstituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclicring, wherein one, two, or three atoms in the heterocyclic ring systemare independently nitrogen, oxygen, or sulfur.
 26. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 27. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 28. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 29. The method of claim19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 30. The method of any oneof claims 19-29, wherein at least one instance of R^(B1) is halogen,substituted or unsubstituted alkyl, or substituted or unsubstituted,monocyclic, 5- to 6-membed heteroaryl, wherein one, two, three, or fouratoms in the heteroaryl ring system are independently nitrogen, oxygen,or sulfur.
 31. The method of any one of claims 19-30, wherein at leastone instance of R^(B2) is halogen or substituted or unsubstituted alkyl.32. The method of any one of claims 19-31, wherein o2 is
 0. 33. Themethod of any one of claims 19-32, wherein at least one instance ofR^(B3) is substituted or unsubstituted heteroaryl or —C(═O)N(R^(a))₂, oran instance of R^(B3) and an instance of R^(B7) are joined to form asubstituted or unsubstituted, monocyclic, 5- to 6-membed heterocyclicring, wherein one, two, or three atoms in the heterocyclic ring systemare independently nitrogen, oxygen, or sulfur.
 34. The method of any oneof claims 19-33, wherein each instance of R^(B4) is hydrogen.
 35. Themethod of any one of claims 19-34, wherein R^(B5) is hydrogen.
 36. Themethod of claim 19, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 37. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (C):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: Ring C1 is a substituted or unsubstituted, monocyclic, 5- to6-membered heterocyclic ring or a substituted or unsubstituted,monocyclic, 5- to 6-membered heteroaryl ring, wherein one or two atomsin the heterocyclic or heteroaryl ring system are nitrogen; Ring C2 is asubstituted or unsubstituted, monocyclic, 6-membered carbocyclic ring ora substituted or unsubstituted phenyl ring; Ring C3 is a substituted orunsubstituted, monocyclic, 5- to 6-membered heterocyclic ring or asubstituted or unsubstituted, monocyclic, 5- to 6-membered heteroarylring, wherein one or two atoms in the heterocyclic or heteroaryl ringsystem are nitrogen; each instance of R^(C1) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; p1 is 0, 1, 2, 3, 4, 5,6, 7, 8, or 9; each instance of R^(C2) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂,or two instances of R^(C2) are joined to form a substituted orunsubstituted carbocyclic ring; p2 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,or 11; each instance of R^(C3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and p3 is 0, 1, 2, 3,4, 5, 6, 7, 8, 9, or
 10. 38. The method of claim 37, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof.
 39. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein Ring C4 is asubstituted or unsubstituted, monocyclic, 5- to 6-membed carbocyclicring.
 40. The method of claim 37, wherein the compound is of theformula:

or a pharmaceutically acceptable salt thereof.
 41. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 42. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 43. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein Ring C5 is asubstituted or unsubstituted, monocyclic, 5- to 6-membed carbocyclicring.
 44. The method of claim 37, wherein the compound is of theformula:

or a pharmaceutically acceptable salt thereof.
 45. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 46. The method of claim37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 47. The method of any oneof claims 37-46, wherein at least one instance of R^(C1) is substitutedor unsubstituted alkyl, —OR^(a), —N(R^(a))₂, or —CN.
 48. The method ofany one of claims 37-47, wherein at least one instance of R^(C2) ishalogen or —OR^(a).
 49. The method of any one of claims 37-48, whereinat least one instance of R^(C3) is substituted or unsubstituted alkyl.50. The method of any one of claims 37-49, wherein p3 is
 0. 51. Themethod of claim 37, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 52. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (D):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: L^(D1) is a substituted or unsubstituted, saturated orunsaturated, C₃₋₇ aliphatic chain, optionally wherein one, two, or threechain atoms of the aliphatic chain are independently replaced with —O—,—S—, —NR^(D6)—, —N═, or ═N—, wherein each instance of R^(D6) isindependently hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; each instance of R^(D1) is independentlyhalogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; q1 is 0, 1, 2, 3, 4, or5; each instance of R^(D2) is independently hydrogen, halogen, orsubstituted or unsubstituted C₁₋₆ alkyl; each instance of R^(D3) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a),—C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; q3 is 0, 1, 2, or 3; each instance of R^(D4) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a),—C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; q4 is 0, 1, 2, or 3; and each instance of R^(D5) isindependently hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group.
 53. The method of claim 52, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein R^(D7) issubstituted or unsubstituted phenyl.
 54. The method of any one of claims52-53, wherein L^(D1) is a substituted or unsubstituted, saturated orunsaturated, C₅ aliphatic chain, optionally wherein one, two, or threechain atoms of the aliphatic chain are independently replaced with —O—,—S—, —NR^(D6)—, ═N—, or —N═.
 55. The method of any one of claims 52-54,wherein at least one instance of R^(D1) is halogen.
 56. The method ofany one of claims 52-55, wherein each instance of R^(D2) is hydrogen.57. The method of any one of claims 52-56, wherein q3 is
 0. 58. Themethod of any one of claims 52-57, wherein q4 is
 0. 59. The method ofany one of claims 52-58, wherein each instance of R^(D5) is hydrogen.60. The method of any one of claims 52-59, wherein at least one instanceof R^(D6) is hydrogen.
 61. The method of claim 52, wherein the compoundis of the formula:

or a pharmaceutically acceptable salt thereof.
 62. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (E):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of culturing step;wherein: X^(E1) is —O—, —S—, —C(═O)—, —C(═S)—, or —C(═NR^(E4))—, whereinR^(E) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; X^(E2) is —O—, —S—, —C(═O)—, or —C(—S)—;R^(E1) is hydrogen, halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂, or R^(E1) and R^(E) are joined to form asubstituted or unsubstituted heterocyclic ring or a substituted orunsubstituted heteroaryl ring; each instance of R^(a) is independentlyhydrogen, substituted or unsubstituted acyl, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, a nitrogen protectinggroup when attached to a nitrogen atom, an oxygen protecting group whenattached to an oxygen atom, or a sulfur protecting group when attachedto a sulfur atom, or two instances of R^(a) are joined to form asubstituted or unsubstituted heterocyclic or substituted orunsubstituted heteroaryl ring; each instance of R^(E2) is independentlyhalogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;r2 is 0, 1, 2, or 3; each instance of R^(E3) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;and r3 is 0, 1, 2, 3, or
 4. 63. The method of claim 62, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof.
 64. The method of any oneof claims 62-63, wherein R^(E1) is substituted or unsubstituted,monocyclic, 5- to 6-membered heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur.
 65. The method of claim 62, wherein the compound isof the formula:

or a pharmaceutically acceptable salt thereof, wherein: each instance ofR^(E5) is independently halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; and r5 is 0, 1, 2, or
 3. 66. The method of claim62, wherein the compound is of the formula:


67. The method of any one of claims 65-66, wherein at least one instanceof R^(E5) is substituted or unsubstituted, monocyclic, 5- to 6-membedheterocyclyl, wherein one, two, or three atoms in the heterocyclic ringsystem are independently nitrogen, oxygen, or sulfur.
 68. The method ofclaim 62, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein X^(E3) is C or N,and Ring E is a substituted or unsubstituted, monocyclic, 5- to6-membered heterocyclic ring or a substituted or unsubstituted,monocyclic, 5- to 6-membered heteroaryl ring, wherein one, two, or threeatoms in the heterocyclic or heteroaryl ring system are independentlynitrogen, oxygen, or sulfur.
 69. The method of claim 62, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein R^(E6) ishydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group.
 70. The method of claim 69, wherein R^(E6) ishydrogen.
 71. The method of any one of claims 62-70, wherein r2 is 0.72. The method of any one of claims 62-71, wherein r3 is
 0. 73. Themethod of claim 62, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 74. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (F):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: X^(F) is —N(R^(F9))—, ═N—, —C(R^(F10))₂—, or ═C(RF¹⁰)—,wherein: R^(F9) is hydrogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, or a nitrogen protecting group;and each instance of R^(F10) is independently hydrogen, halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; R^(F1) is absent,hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group; R^(F2) is hydrogen, halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;R^(F3) is absent, hydrogen, halogen, or substituted or unsubstitutedC₁₋₆ alkyl, or R^(F3) and R^(F2) are joined to form ═O; R^(F4) ishydrogen, halogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;R^(F5) is absent, hydrogen, halogen, or substituted or unsubstitutedC₁₋₆ alkyl; R^(F6) is hydrogen, halogen, substituted or unsubstitutedalkyl, substituted or unsubstituted alkenyl, substituted orunsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; R^(F7) is hydrogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group; and R^(F8) is hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group.75. The method of claim 74, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 76. The method of claim74, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 77. The method of claim74, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 78. The method of any oneof claims 74-77, wherein R^(F1) is absent, substituted or unsubstitutedalky, or substituted or unsubstituted, monocyclic, 5- to 6-membedcarbocyclyl.
 79. The method of any one of claims 74-78, wherein R^(F2)is —NR^(a)C(═O)N(R^(a))₂.
 80. The method of any one of claims 74-78,wherein R^(F2) and R^(F3) are joined to form ═O.
 81. The method of anyone of claims 74-80, wherein R^(F4) is hydrogen or substituted orunsubstituted alkyl.
 82. The method of any one of claims 74-81, whereinR^(F5) is absent or hydrogen.
 83. The method of any one of claims 74-82,wherein R^(F6) is hydrogen.
 84. The method of any one of claims 74-83,wherein R^(F7) is substituted or unsubstituted alkyl or substituted orunsubstituted, monocyclic, 5- to 6-membed heteroaryl, wherein one, two,three, or four atoms in the heteroaryl ring system are independentlynitrogen, oxygen, or sulfur.
 85. The method of any one of claims 74-84,wherein R^(F8) is hydrogen.
 86. The method of any one of claims 74-85,wherein R^(F9) is substituted or unsubstituted phenyl.
 87. The method ofany one of claims 74-86, wherein at least one instance of R^(F10) issubstituted or unsubstituted phenyl, or —C(═O)R^(a).
 88. The method ofclaim 74, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 89. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (G):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of culturing step;wherein: Ring G is a substituted or unsubstituted phenyl ring or asubstituted or unsubstituted, monocyclic, 5- to 6-membered heteroarylring, wherein one or two atoms in the heteroaryl ring system areindependently nitrogen, oxygen, or sulfur; each instance of R^(G1) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a),—C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; each instance of R^(a) is independently hydrogen,substituted or unsubstituted acyl, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, a nitrogen protecting groupwhen attached to a nitrogen atom, an oxygen protecting group whenattached to an oxygen atom, or a sulfur protecting group when attachedto a sulfur atom, or two instances of R^(a) are joined to form asubstituted or unsubstituted heterocyclic or substituted orunsubstituted heteroaryl ring; t1 is 0, 1, 2, 3, 4, or 5; L^(G) is abond, —O—, —S—, —N(R^(G5))—, or —C(R^(G6))₂—, wherein R^(G5) ishydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; and each instance of R^(G6) is independently hydrogen,halogen, or substituted or unsubstituted C₁₋₆ alkyl; R^(G2) is hydrogen,halogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, or substituted or unsubstitutedheteroaryl; R^(G3) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl,or a nitrogen protecting group; each instance of R^(G4) is independentlyhalogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), —OC(═O)N(R^(a))₂,—S(═O)₂R^(a), —S(═O)₂OR^(a), or —S(═O)₂N(R^(a))₂; and t4 is 0, 1, 2, 3,or
 4. 90. The method of claim 89, wherein the compound is of theformula:

or a pharmaceutically acceptable salt thereof.
 91. The method of claim89, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 92. The method of claim89, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 93. The method of any oneof claims 89-92, wherein at least one instance of R^(G1) is substitutedor unsubstituted alkyl, —C(═O)N(R^(a))₂, or —N(R^(a))C(═O)R^(a).
 94. Themethod of any one of claims 89-93, wherein R^(G2) is substituted orunsubstituted phenyl.
 95. The method of any one of claims 89-94, whereinR^(G3) is hydrogen.
 96. The method of any one of claims 89-95, whereinat least one instance of R^(G4) is halogen, —C(═O)OR^(a), —S(═O)₂R^(a),or —S(═O)₂N(R^(a))₂.
 97. The method of any one of claims 89-96, whereinR^(G5) is hydrogen.
 98. The method of claim 89, wherein the compound isof the formula:

or a pharmaceutically acceptable salt thereof.
 99. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (H):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: L^(H) is a substituted or unsubstituted, saturated orunsaturated, C₁₋₄ aliphatic chain, optionally wherein one or two chainatoms of the aliphatic chain are independently replaced with —O—, —S—,—NR^(H5)—, —N═, or ═N—, wherein each instance of R^(H5) is independentlyhydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, or a nitrogen protecting group; each instance of R^(H1) isindependently halogen, substituted or unsubstituted alkyl, substitutedor unsubstituted alkenyl, substituted or unsubstituted alkynyl,substituted or unsubstituted carbocyclyl, substituted or unsubstitutedheterocyclyl, substituted or unsubstituted aryl, substituted orunsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN,—C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a),—C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; each instance of R^(a) is independently hydrogen,substituted or unsubstituted acyl, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, a nitrogen protecting groupwhen attached to a nitrogen atom, an oxygen protecting group whenattached to an oxygen atom, or a sulfur protecting group when attachedto a sulfur atom, or two instances of R^(a) are joined to form asubstituted or unsubstituted heterocyclic or substituted orunsubstituted heteroaryl ring; u1 is 0, 1, 2, 3, 4, or 5; R^(H2) ishydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; each instance of R^(H3) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;u3 is 0, 1, or 2; each instance of R^(H4) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;and u4 is 0, 1, 2, 3, 4, or
 5. 100. The method of claim 99, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof.
 101. The method of claim99, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 102. The method of claim99, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 103. The method of anyone of claims 99-102, wherein at least one instance of R^(H1) issubstituted or unsubstituted alkyl or substituted or unsubstituted,monocyclic, 5- to 6-membed heterocyclyl, wherein one, two, or threeatoms in the heterocyclic ring system are independently nitrogen,oxygen, or sulfur.
 104. The method of any one of claims 99-103, whereinR^(H2) is hydrogen.
 105. The method of any one of claims 99-104, whereinat least one instance of R^(H3) is halogen.
 106. The method of any oneof claims 99-105, wherein u3 is
 0. 107. The method of any one of claims99-106, wherein at least one instance of R^(H4) is substituted orunsubstituted alkyl, or —C(═O)N(R^(a))₂.
 108. The method of any one ofclaims 99-107, wherein at least one instance of R^(H5) is hydrogen orsubstituted or unsubstituted phenyl.
 109. The method of claim 99,wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 110. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (I):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: R^(I1) is substituted or unsubstituted alkyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, or substituted or unsubstitutedheteroaryl; R^(I2) is hydrogen or substituted or unsubstituted alkyl;R^(I5) is hydrogen, substituted or unsubstituted alkyl, substituted orunsubstituted aralkyl, or substituted or unsubstituted carbocyclyl;R^(I6) is hydrogen or substituted or unsubstituted alkyl; each instanceof R^(I7) is independently halogen, substituted or unsubstituted alkyl,substituted or unsubstituted aralkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —O-(substituted orunsubstituted alkyl), —NH-(substituted or unsubstituted alkyl),—NH-(substituted or unsubstituted aryl), —N(substituted or unsubstitutedalkyl)-(substituted or unsubstituted alkyl), —N(substituted orunsubstituted alkyl)-(substituted or unsubstituted aryl), —NO₂, or —CN;and v7 is 0, 1, 2, 3, or
 4. 111. The method of claim 110, wherein thecompound is of the formula:

or a pharmaceutically acceptable salt thereof, wherein: each instance ofR^(I8) is independently halogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a),—CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂,—C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a),—NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a),or —OC(═O)N(R^(a))₂; each instance of R^(a) is independently hydrogen,substituted or unsubstituted acyl, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl,substituted or unsubstituted heteroaryl, a nitrogen protecting groupwhen attached to a nitrogen atom, an oxygen protecting group whenattached to an oxygen atom, or a sulfur protecting group when attachedto a sulfur atom, or two instances of R^(a) are joined to form asubstituted or unsubstituted heterocyclic or substituted orunsubstituted heteroaryl ring; and v8 is 0, 1, 2, 3, 4, or
 5. 112. Themethod of claim 111, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 113. The method of anyone of claims 110-112, wherein R^(I2) is substituted or unsubstitutedalkyl.
 114. The method of any one of claims 110-113, wherein R^(I5) issubstituted or unsubstituted alkyl.
 115. The method of any one of claims110-114, wherein R^(I6) is hydrogen.
 116. The method of any one ofclaims 110-115, wherein v7 is
 0. 117. The method of any one of claims111-116, wherein v8 is
 0. 118. The method of any one of claims 111-117,wherein R^(a) is substituted or unsubstituted C₁₋₆ alkyl.
 119. Themethod of claim 110, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 120. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (J):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: L^(J) is a substituted or unsubstituted, saturated orunsaturated, C₁₋₆ aliphatic chain, optionally wherein one or two chainatoms of the aliphatic chain are independently replaced with —O—, —S—,—NR^(J6)—, —N═, or ═N—, wherein each instance of R^(J6) is independentlyhydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; each instance of R^(J1) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; w1 is 0, 1, 2, 3, or 4;R^(J2) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; R^(J3) is hydrogen, halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; R^(J4) is hydrogen,substituted or unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group;each instance of R^(J5) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; and w5 is 0, 1, 2, 3,4, or
 5. 121. The method of claim 120, wherein L^(J) is a substituted orunsubstituted, saturated or unsaturated, C₃ aliphatic chain, optionallywherein one or two chain atoms of the aliphatic chain are independentlyreplaced with —O—, —S—, —NR^(J6)—, —N═, or ═N—.
 122. The method of claim120, wherein L^(J) is of the formula:

wherein each instance of R^(J7) is independently hydrogen, halogen, orsubstituted or unsubstituted C₁₋₆ alkyl.
 123. The method of claim 120,wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 124. The method of anyone of claims 120-123, wherein w1 is
 0. 125. The method of any one ofclaims 120-124, wherein R^(J2) is hydrogen.
 126. The method of any oneof claims 120-125, wherein R^(J3) is substituted or unsubstituted alkyl.127. The method of any one of claims 120-126, wherein R^(J4) issubstituted or unsubstituted alkyl.
 128. The method of any one of claims120-127, wherein at least one instance of R^(J5) is halogen.
 129. Themethod of any one of claims 120-128, wherein at least one instance ofR^(J6) is hydrogen.
 130. The method of claim 120, wherein the compoundis of the formula:

or a pharmaceutically acceptable salt thereof.
 131. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (K):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: R^(K1) is hydrogen, substituted or unsubstituted alkyl,substituted or unsubstituted alkenyl, substituted or unsubstitutedalkynyl, substituted or unsubstituted carbocyclyl, substituted orunsubstituted heterocyclyl, substituted or unsubstituted aryl, orsubstituted or unsubstituted heteroaryl; R^(K2) is hydrogen, substitutedor unsubstituted C₁₋₆ alkyl, or a nitrogen protecting group; eachinstance of R^(K3) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; each instance of R^(a)is independently hydrogen, substituted or unsubstituted acyl,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; x3 is 0, 1, 2, 3, or 4;R^(K4) is hydrogen, substituted or unsubstituted C₁₋₆ alkyl, or anitrogen protecting group; each instance of R^(K5) is independentlyhalogen, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;and x5 is 0, 1, 2, 3, 4, or
 5. 132. The method of claim 131, whereinR^(K)1 is substituted or unsubstituted alkyl.
 133. The method of claim131, wherein R^(K1) is C₁₋₆ alkyl substituted with at least one —OH.134. The method of any one of claims 131-133, wherein R^(K2) ishydrogen.
 135. The method of any one of claims 131-134, wherein at leastone instance of R^(K3) is halogen.
 136. The method of any one of claims131-135, wherein R^(K4) is hydrogen.
 137. The method of any one ofclaims 131-136, wherein at least one instance of R^(K5) is halogen. 138.The method of claim 131, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 139. A method forconverting pluripotency state of a vertebrate cell to a more naïvestate, the method comprising: culturing a pluripotent vertebrate cell inthe presence of a compound of Formula (L):

or a pharmaceutically acceptable salt thereof; and maintaining the cellin culture under conditions suitable and a time sufficient to convertthe pluripotency state of the vertebrate cell to a more naïve state thanthe pluripotency state of the vertebrate cell of the step of culturing;wherein: L^(L) is a substituted or unsubstituted, saturated orunsaturated, C₁₋₄ aliphatic chain, optionally wherein one or two chainatoms of the aliphatic chain are independently replaced with —O—, —S—,—NR^(L5)—, —N═, or ═N—, wherein each instance of R^(L5) is independentlyhydrogen, substituted or unsubstituted C₁₋₆ alkyl, or a nitrogenprotecting group; each instance of R^(L1) is independently halogen,substituted or unsubstituted alkyl, substituted or unsubstitutedalkenyl, substituted or unsubstituted alkynyl, substituted orunsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, —OR^(a), —N(R^(a))₂, —SR^(a), —CN, —SCN, —C(═NR^(a))R^(a),—C(═NR^(a))OR^(a), —C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a),—C(═O)N(R^(a))₂, —NO₂, —NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a),—NR^(a)C(═O)N(R^(a))₂, —OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂;each instance of R^(a) is independently hydrogen, substituted orunsubstituted acyl, substituted or unsubstituted alkyl, substituted orunsubstituted alkenyl, substituted or unsubstituted alkynyl, substitutedor unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,substituted or unsubstituted aryl, substituted or unsubstitutedheteroaryl, a nitrogen protecting group when attached to a nitrogenatom, an oxygen protecting group when attached to an oxygen atom, or asulfur protecting group when attached to a sulfur atom, or two instancesof R^(a) are joined to form a substituted or unsubstituted heterocyclicor substituted or unsubstituted heteroaryl ring; y1 is 0, 1, 2, 3, or 4;each instance of R^(L2) is independently halogen, substituted orunsubstituted alkyl, substituted or unsubstituted alkenyl, substitutedor unsubstituted alkynyl, substituted or unsubstituted carbocyclyl,substituted or unsubstituted heterocyclyl, substituted or unsubstitutedaryl, substituted or unsubstituted heteroaryl, —OR^(a), —N(R^(a))₂,—SR^(a), —CN, —SCN, —C(═NR^(a))R^(a), —C(═NR^(a))OR^(a),—C(═NR^(a))N(R^(a))₂, —C(═O)R^(a), —C(═O)OR^(a), —C(═O)N(R^(a))₂, —NO₂,—NR^(a)C(═O)R^(a), —NR^(a)C(═O)OR^(a), —NR^(a)C(═O)N(R^(a))₂,—OC(═O)R^(a), —OC(═O)OR^(a), or —OC(═O)N(R^(a))₂; y2 is 0, 1, 2, 3, 4,5, 6, 7, 8, 9, or 10; each instance of R^(L3) is independently hydrogen,halogen, or substituted or unsubstituted C₁₋₆ alkyl; and each instanceof R^(L4) is independently hydrogen, substituted or unsubstituted C₁₋₆alkyl, or a nitrogen protecting group.
 140. The method of claim 139,wherein L^(L) is a substituted or unsubstituted, saturated orunsaturated, C₂ aliphatic chain, optionally wherein one chain atom ofthe aliphatic chain is replaced with —O—, —S—, —NR^(L)—, ═N—, or —N═.141. The method of claim 139, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 142. The method of claim139, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 143. The method of anyone of claims 139-142, wherein y1 is
 0. 144. The method of any one ofclaims 139-143, wherein y2 is
 0. 145. The method of any one of claims139-144, wherein each instance of R^(L3) is independently hydrogen orsubstituted or unsubstituted alkyl.
 146. The method of any one of claims139-145, wherein each instance of R^(L4) is hydrogen.
 147. The method ofclaim 139, wherein the compound is of the formula:

or a pharmaceutically acceptable salt thereof.
 148. The method of anyone of claims 1-147, wherein the culture comprises fibroblast growthfactor and/or Activin A.
 149. The method of any one of claims 1-148,wherein the vertebrate cell is a human stem cell.
 150. The method ofclaim 149, wherein the human stem cell is a human embryonic stem cell.151. The method of any one of claims 1-148, wherein the vertebrate cellis a human induced pluripotent cell (iPS cell).
 152. The method of anyone of claims 1-148, wherein the vertebrate cell is derived from a humanembryonic stem cell line.
 153. The method of any one of claims 1-148,wherein the vertebrate cell is derived from an embryo.
 154. The methodof any one of claims 1-153, wherein the more naïve pluripotentvertebrate cell of the step of maintaining uses the distal Oct4 enhancerelement for OCT4 expression.
 155. The method of any one of claims 1-154,wherein the more naïve pluripotent vertebrate cell of the step ofmaintaining has a global gene expression profile which clusters withnaïve mouse ESCs as opposed to stem cell lines derived from mouseepiblast (EpiSCs) and/or less naïve human ESCs.
 156. A compositioncomprising naïve pluripotent vertebrate cells produced by the methods ofany one of claims 1-155.
 157. A naïve pluripotent vertebrate cell lineproduced by the methods of any one of claims 1-155.
 158. A naïvepluripotent vertebrate cell, wherein the cell uses the distal Oct4enhancer element for OCT4 expression.
 159. The naïve pluripotentvertebrate cell of claim 158, wherein the cell is a primate cell andwherein the cell uses the endogenous distal OCT4 enhancer element forOct4 expression.
 160. The naïve pluripotent vertebrate cell of claim158, wherein the cell is a human cell and wherein the cell uses theendogenous distal OCT4 enhancer element for Oct4 expression.
 161. Anaïve pluripotent vertebrate cell, wherein the cell has a global geneexpression profile which clusters with naïve mouse ESCs as opposed tostem cell lines derived from mouse epiblast (EpiSCs) and/or less naïvehuman ESCs.
 162. The naïve pluripotent vertebrate cell of claim 161,wherein the cell is a primate cell.
 163. The naïve pluripotentvertebrate cell of claim 161, wherein the cell is a human cell.
 164. Thenaïve pluripotent vertebrate cell of any one of claims 158 to 163,wherein the cell is produced by the methods of any one of claims 1-155.165. A kit for culturing vertebrate cells, the kit comprising: aserine/threonine-protein kinase B-Raf (BRAF) inhibitor, an epidermalgrowth factor receptor (EGFR) inhibitor, a vascular endothelial growthfactor 1 (VEGFR1) inhibitor, or a fibroblast growth factor receptor 1(FGFR1) inhibitor; and instructions for culturing vertebrate cells. 166.The kit of claim 165, wherein the kit further comprisesmitogen-activated protein kinase kinase (MEK) inhibitor.
 167. The kit ofclaims 165 or 166, wherein the kit further comprises a glycogen synthasekinase 3 (GSK3) inhibitor, a rho-associated protein kinase (ROCK)inhibitor, and/or a proto-oncogene tyrosine-protein kinase (Src)inhibitor.
 168. The kit of any one of claims 165-167, wherein the kitcomprises a BRAF inhibitor, a MEK inhibitor, and a GSK3 inhibitor. 169.The kit of any one of claims 165-167, wherein the kit comprises a BRAFinhibitor, a MEK inhibitor, a GSK3 inhibitor, and a ROCK inhibitor. 170.The kit of any one of claims 165-167, wherein the kit comprises a BRAFinhibitor, a MEK inhibitor, a GSK3 inhibitor, a ROCK inhibitor, and anSrc inhibitor.
 171. The kit of any one of claims 165-167, wherein thekit comprises a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, andan Src inhibitor.
 172. The kit of any one of claims 165-171, wherein thekit further comprises cell culture medium.
 173. The kit of any one ofclaims 165-172, wherein the vertebrate cells are naïve pluripotentcells, human pluripotent cells, or naïve human pluripotent cells.
 174. Akit for preparing a cell culture medium, the kit comprising: aserine/threonine-protein kinase B-Raf (BRAF) inhibitor, an epidermalgrowth factor receptor (EGFR) inhibitor, a vascular endothelial growthfactor 1 (VEGFR1) inhibitor, or a fibroblast growth factor receptor 1(FGFR1) inhibitor; and instructions for preparing a cell culture medium.175. The kit of claim 174, wherein the kit further comprisesmitogen-activated protein kinase kinase (MEK) inhibitor.
 176. The kit ofclaims 174 or 175, wherein the kit further comprises a glycogen synthasekinase 3 (GSK3) inhibitor, a rho-associated protein kinase (ROCK)inhibitor, and/or a proto-oncogene tyrosine-protein kinase (Src)inhibitor.
 177. The kit of any one of claims 174-176, wherein the kitcomprises a BRAF inhibitor, a MEK inhibitor, and a GSK3 inhibitor. 178.The kit of any one of claims 174-176, wherein the kit comprises a BRAFinhibitor, a MEK inhibitor, a GSK3 inhibitor, and a ROCK inhibitor. 179.The kit of any one of claims 174-176, wherein the kit comprises a BRAFinhibitor, a MEK inhibitor, a GSK3 inhibitor, a ROCK inhibitor, and anSrc inhibitor.
 180. The kit of any one of claims 174-176, wherein thekit comprises a BRAF inhibitor, a MEK inhibitor, a ROCK inhibitor, andan Src inhibitor.
 181. The kit of any one of claims 174-180, wherein thekit further comprises a basal medium.
 182. The kit of any one of claims174-181, wherein the basal medium is a serum-free medium.
 183. The kitof any one of claims 174-182, wherein the basal medium comprises one ormore supplements.
 184. The method of any one of claims 1-155 or the kitof any one of claims 165-183, wherein the BRAF inhibitor is of theformula:

or a pharmaceutically acceptable salt thereof.
 185. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theVEGFR1 inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 186. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theFGFR1 inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 187. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theMEK inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 188. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theGSK3 inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 189. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theROCK inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 190. The method of anyone of claims 1-155 or the kit of any one of claims 165-183, wherein theSrc inhibitor is of the formula:

or a pharmaceutically acceptable salt thereof.
 191. A cell culturemedium comprising: a basal medium; and a serine/threonine-protein kinaseB-Raf (BRAF) inhibitor, an epidermal growth factor receptor (EGFR)inhibitor, a vascular endothelial growth factor 1 (VEGFR1) inhibitor, ora fibroblast growth factor receptor 1 (FGFR1) inhibitor.
 192. The cellculture medium of claim 191, wherein the cell culture medium furthercomprises mitogen-activated protein kinase kinase (MEK) inhibitor. 193.The cell culture medium of claim 191 or 192, wherein the cell culturemedium further comprises a glycogen synthase kinase 3 (GSK3) inhibitor,a rho-associated protein kinase (ROCK) inhibitor, and/or aproto-oncogene tyrosine-protein kinase (Src) inhibitor.
 194. The cellculture medium of any one of claims 191-193, wherein the cell culturemedium comprises a BRAF inhibitor, a MEK inhibitor, and a GSK3inhibitor.
 195. The cell culture medium of any one of claims 191-193,wherein the cell culture medium comprises a BRAF inhibitor, a MEKinhibitor, a GSK3 inhibitor, and a ROCK inhibitor.
 196. The cell culturemedium of any one of claims 191-193, wherein the cell culture mediumcomprises a BRAF inhibitor, a MEK inhibitor, a GSK3 inhibitor, a ROCKinhibitor, and an Src inhibitor.
 197. The cell culture medium of any oneof claims 191-193, wherein the cell culture medium comprises a BRAFinhibitor, a MEK inhibitor, a ROCK inhibitor, and an Src inhibitor. 198.The cell culture medium of any one of claims 191-197, wherein the basalmedium is a serum-free medium.
 199. The cell culture medium of any oneof claims 191-198, wherein the basal medium comprises one or moresupplements.